Em Barratt

Mating patterns, relatedness and the basis of natal philopatry in the brown long‐eared bat, Plecotus auritus

The brown long‐eared bat, Plecotus auritus, is unusual among temperate zone bats in that summer maternity colonies are composed of adult males and females, with both sexes displaying natal philopatry and long‐term association with a colony. Here, we describe the use of microsatellite analysis to investigate colony relatedness and mating patterns, with the aim of identifying the evolutionary determinants of social organization in P. auritus. Mean colony relatedness was found to be low (R = 0.033 ± 0.002), with pairwise estimates of R within colonies ranging from −0.4 to 0.9. The proportion of young fathered by males in their own colony was investigated using a Bayesian approach, incorporating parameters detailing the number of untyped individuals. This analysis revealed that most offspring were fathered by males originating from a different colony to their own. In addition, we determined that the number of paternal half‐sibs among cohorts of young was low, inferring little or no skew in male reproductive success. The results of this study suggest that kin selection cannot account for colony stability and natal philopatry in P. auritus, which may instead be explained by advantages accrued through the use of familiar and successful roost sites, and through long‐term associations with conspecifics. Moreover, because the underlying causes of male natal dispersal in mammals, such as risk of inbreeding or competition for mates, appear to be avoided via extra‐colony copulation and low male reproductive skew, both P. auritus males and females are able to benefit from long‐term association with the natal colony.

Genetic variation and population structure in the endangered greater horseshoe bat Rhinolophus ferrumequinum

Following a dramatic decline last century, the British population of the endangered greater horseshoe bat Rhinolophus ferrumequinum is highly fragmented. To examine the consequences of fragmentation and limited dispersal on patterns of genetic structure and variation, we used microsatellite markers to screen bats from around 50% of the known maternity colonies in Britain, and two areas from continental Europe. Analyses revealed that Welsh and English colonies were genetically isolated. This, and lower variability in Britain than north France, may result from either genetic drift, or the species’ colonization history. Gene flow among most neighbouring colonies was not generally restricted, with one exception. These findings have important implications for the ongoing conservation management of this species.

Outbreeding increases offspring survival in wild greater horseshoe bats (Rhinolophus ferrumequinum)

The factors influencing the survival of greater horseshoe bat (Rhinolophus ferrumequinum) offspring born over seven years at a maternity colony in south-west Britain were studied. The effects of a range of phenotypic and maternal variables were analysed using a historical data set. In addition, the influence of two genetic measures on mortality, individual heterozygosity and a new measure of outbreeding, termed mean d2, was assessed. Logistic regressions were undertaken with survival modelled as a binary response variable. Survival to two life stages was studied for each variable and all models were developed for both sexes separately and together. Only one variable, mean d2, was significantly associated with survival. Male offspring with high mean d2 scores were more likely to survive to their first and second summers. The influence of mean d2 was not due to a single locus under selection but a wider multilocus effect and probably represents heterosis as opposed to solely inbreeding depression. Therefore, the extent to which an individual is outbred may determine survival more than widely used phenotypic characteristics such as size and mass. Mean d2 may reflect immunocompetence, which influences mortality. Protection of mating sites in order to facilitate gene flow and, therefore, outbreeding may help to promote population stability and growth.

Parentage, reproductive success and breeding behaviour in the greater horseshoe bat (Rhinolophus ferrumequinum)

Female greater horseshoe bats form maternity colonies each summer in order to give birth and raise young. During the mating period, females visit males occupying territorial sites, copulation takes place and sperm are stored until ovulation occurs, normally in April. Using microsatellite markers and a likelihood method of parentage analysis, we studied breeding behaviour and male reproductive success over a five–year period in a population of bats in south–west Britain. Paternity was assigned with 80% confidence to 44% of young born in five successive cohorts. While a small annual skew in male reproductive success was detected, the variance increased over five years due to the repeated success of a few individuals. Mating was polygynous, although some females gave birth to offspring sired by the same male in separate years. Such repeated partnerships probably result from fidelity for either mating sites or individuals or from sperm competition. Females mated with males born both within and outside their own natal colony; however, relatedness between parents was no less than the average recorded for male–female pairs. Gene flow between colonies is likely to be primarily mediated by both female and male dispersal during the mating period rather than more permanent movements.

Genetic structure of fragmented populations of red squirrel (Sciurus vulgaris) in the UK

The relationships among 207 squirrels from 12 locations in the UK and three in mainland Europe were examined using mitochondrial DNA (mtDNA) control region sequence. Twenty‐six haplotypes were detected, many of which were population specific. Eighty per cent of the populations analysed contained two or more haplotypes. Hierarchical analysis of molecular variance showed the majority of genetic variation to be partitioned among populations. Genetic diversity varied considerably within the UK, and conformed to no obvious geographical trend. The populations in Argyll and Spadeadam Forest showed the highest levels of variation in the UK. However, the greatest genetic diversity was seen in Bavaria, southern Germany where six unique alleles were detected in a sample of 10 individuals. Phylogenetic analysis revealed no evolutionary divergence between UK and mainland European haplotypes. We conclude that, within the UK, the genetic patterns observed are most likely to be explained by the effects of genetic drift which has occurred since the isolation of populations during the past few hundred years, hence we cannot detect any underlying phylogeographic pattern. Therefore, the use of larger, geographically distinct populations within the UK for augmentation of small isolated populations is unlikely to pose problems of genetic incompatibility. Further, the role that demographic factors may have in complicating the application of current genetically based management unit criteria is likely to need further attention.

Can skull morphology be used to predict ecological relationships between bat species? A test using two cryptic species of pipistrelle

Can ecological relationships between bat species be predicted largely on the basis of morphology? This question was addressed by investigating skull morphology of two cryptic species of the pipistrelle bat. Since 45 P. pipistrellus apparently eats larger prey than 55 P. pipistrellus, we predicted that it would have a larger overall skull size, a larger dentary apparatus, and a larger gape. To test these predictions, variables were measured from skulls of the two cryptic species, and comparisons made between them. In accordance with our predictions, overall skull size was larger in 45 P. pipistrellus than in 55 P. pipistrellus, and 45 P. pipistrellus had a longer lower jaw and the distance between the jaws at maximum gape was larger. In addition, 45 P. pipistrellus had longer upper canines, which may allow it to pierce harder prey items than 55 P. pipistrellus. Only some aspects of dietary differences between the two cryptic species could be explained by differences in skull morphology, and we suggest that empirical data, at least on diet and habitat use, are also required to explain mechanisms of resource partitioning among species in bat communities.

Characterization of microsatellite loci in the greater horseshoe bat Rhinolophus ferrumequinum

A full understanding of colony organization and life histories in social insects can only be achieved by investigating the genetic structure of colonies and populations (Pamilo et al . 1997). Although most genetic studies on ants have been performed on species with complex colony organization (but see Tay et al . 1997), the study of morphologically primitive ants, characterized by limited caste dimorphism and small colony size, could provide new insights into the evolution of sociality (Peeters 1997). In some morphologically primitive species of Ponerine ants, the queen caste has been lost and one worker mates (the gamergate) and produces diploid offspring (Peeters 1991). In these queenless species the foundation of new colonies occurs only by fission. Even in such a relatively simple social organization, polymorphic genetic markers will be required to estimate the rate of gamergate turnover and colony fission, the extent of population viscosity and their effects on the evolution of sociality. In this note, we characterize eight polymorphic microsatellite markers from the queenless species Diacamma cyaneiventre . Microsatellite markers have been characterized in a number of groups of ants (e.g. Chapuisat 1996; Herbers & Mouser 1998), but none are available for ponerine ants. We also report results of cross-amplification on 11 Ponerine species (seven Diacamma species and four belonging to other genera of Ponerinae). Genomic DNA from nine Diacamma cyaneiventre larvae was extracted by a high-salt procedure using NaCl and digested with the BSP 143I restriction enzyme. Fragments between 300 and 600 bp were selected and ligated to a Bam HIdigested pBluescript II KS+ vector (Stratagene) and cloned in Escherichia coli SL-1 Blue cells (Stratagene). Synthetic oligonucleotides (TC) 10 and (TG) 10 , labelled with the DIG system (Boehringer Mannheim) were used to screen about 1200 recombinant colonies. Out of 146 positive clones, 78 were purified with Qiaprep (Qiagen) and 24 were sequenced either manually using the T7-sequencing kit (Pharmacia) or an ALF express automatic sequencer (Pharmacia). Primers flanking microsatellite repeats were designed for 10 loci using Primer 3 software (Rozen & Skaletsky 1996). Genomic DNA was prepared following either a classic phenol/ chloroform or high-salt extraction and diluted to 1/5 before amplification. PCR reaction mixtures (10 μ L final volume) contained 50 ng of template DNA, 75 μ m dCTP, GTP and dTTP, 7.5 μ m dATP, 0.025 μ Ci 33 P-dATP (Amersham), 4 pmoles each primer, 1 × Taq buffer (containing 1.5 m m MgCl 2 ) and 0.25 U Taq DNA polymerase (Qiagen). For the D18 locus, the amount of dNTP, 33 P-dATP and Taq DNA polymerase was doubled to allow a better amplification of the longest allele in heterozygote individuals. Amplifications were performed in a PTC-100 (MJ Research) thermal cycler using the following parameters: 3 min at 94 ° C followed by 30 cycles with 30 s at 94 ° C, 30 s at the annealing temperature (see Table 1) and 1 min at 72 ° C, and a final elongation step of 10 min at 72 ° C. Amplification products were run on 6% polyacrylamide sequencing gels using an M13 sequence as a size marker. All 10 microsatellite loci gave repeatable and scorable patterns. An estimate of the variability of these loci was determined using a sample of 45 unrelated workers. Eight loci were polymorphic (Table 1). The expected heterozygosity ranged from 0.39 to 0.95, the highest diversity being observed for an uninterrupted microsatellite showing the largest number of repeats (29). No significant deviations from Hardy– Weinberg proportions were detected using a subsample of 33 individuals collected in a locality near Kotigehara (Table 1). Results of the cross-amplification are shown in Table 2. PCR conditions were as described above except that the annealing temperature was 52 ° C for all primers. Where only one individual was extracted, DNA quality was confirmed using mitochondrial DNA primers known to amplify on these species (data not shown). Within the genus Diacamma , cross-species amplification was successful with an average of six pairs of primers amplifying per species. For D. ceylonense and D. sp. (nilgiri) from India, nine individuals were screened to check for polymorphism. Four microsatellites were polymorphic in D. ceylonense and three in D. sp. (nilgiri) . When used on different genera, the efficiency of amplification was low even though these genera belong to the same tribe as Diacamma . Our results may therefore indicate a recent common ancestor for the Diacamma species studied. In conclusion, given their level of polymorphism, the primers presented here should prove to be very useful for investigating population and colony genetic structure of different Diacamma species.