Robert J. Whelan

Patterns of hybridization and asymmetrical gene flow in hybrid zones of the rare Eucalyptus aggregata and common E. rubida

The patterns of hybridization and asymmetrical gene flow among species are important for understanding the processes that maintain distinct species. We examined the potential for asymmetrical gene flow in sympatric populations of Eucalyptus aggregata and Eucalyptus rubida, both long-lived trees of southern Australia. A total of 421 adults from three hybrid zones were genotyped with six microsatellite markers. We used genealogical assignments, admixture analysis and analyses of spatial genetic structure and spatial distribution of individuals, to assess patterns of interspecific gene flow within populations. A high number of admixed individuals were detected (13.9–40% of individuals), with hybrid populations consisting of F1 and F2 hybrids and backcrosses in both parental directions. Across the three sites, admixture proportions were skewed towards the E. aggregata genetic cluster (x=0.56–0.65), indicating that backcrossing towards E. aggregata is more frequent. Estimates of long-term migration rates also indicate asymmetric gene flow, with higher migration rates from E. aggregata to hybrids compared with E. rubida. Taken together, these results indicate a greater genetic input from E. aggregata into the hybrid populations. This asymmetry probably reflects differences in style lengths (E. rubida: ∼7 mm, E. aggregata: ∼4 mm), which can prevent pollen tubes of smaller-flowered species from fertilizing larger-flowered species. However, analyses of fine-scale genetic structure suggest that localized seed dispersal (<40 m) and greater clustering between hybrid and E. aggregata individuals may also contribute to directional gene flow. Our study highlights that floral traits and the spatial distributions of individuals can be useful predictors of the directionality of interspecific gene flow in plant populations.

The birds and the bees: pollinator behaviour and variation in the mating system of the rare shrub Grevillea macleayana

BACKGROUND AND AIMS In Australia, honey-bees have invaded systems that evolved without social insect pollinators, where many plants are adapted to vertebrate pollination. Behavioural differences between pollinators are likely to influence mating patterns, but few studies have examined this empirically in long-lived, woody, perennials. It was shown previously that outcrossing rates in Grevillea macleayana vary among populations. Here tests were conducted to determine whether the behaviour of birds and honey-bees differed between a population previously found to be highly outcrossed and two inbreeding populations. METHODS Visit frequencies and movement patterns of the visitors to inflorescences at three sites over two seasons were compared. A caging experiment was used to test the effects of excluding birds on pollen removal from newly opened flowers and on pollen deposition on stigmas that had been washed clean. KEY RESULTS Honey-bees were the most frequent visitors overall, but honeyeaters were more frequent visitors in the population previously found to have a high outcrossing rate than they were in either of the other populations. More visits by honeyeaters were from distant plants. Pollen removal did not vary greatly among sites, and was not affected by bird exclusion; however, more pollen was deposited on the stigmas of cleaned pollen presenters in the population previously observed to be highly outcrossing than in the other two. This high level of pollen deposition was reduced by experimental bird exclusion. CONCLUSIONS Honey-bees were the most frequent visitors, by an order of magnitude, and excluding vertebrates revealed that bees were removing most of the pollen but deposited fewer pollen grains on stigmas. Birds were more frequent visitors at the site previously found to be outcrossing than the other two sites, and they moved further between plants and visited fewer inflorescences on a plant during a foraging bout than bees did. These characteristics of bird visits to G. macleayana would be sufficient to produce significant variation in outcrossing rates among sites.

Distinguishing between persistence and dormancy in soil seed banks of three shrub species from fire-prone southeastern Australia

Question: Is primary dormancy required for seed bank persistence of Leucopogon species? How does the distinction between dormancy and persistence affect our understanding of seed bank dynamics in fire-prone regions? Location: Sclerophyllous plant communities, Sydney region, southeastern Australia. Methods: Seed bank longevity of three morphophysiologically dormant species was ascertained using seed burial trials. Seeds of each species were buried in situ in replicate mesh bags and retrieved annually over three years. Laboratory germination trials and embryo growth measurements over time were used to establish in which order each component of dormancy was overcome, and whether seed banks persisted after the loss of primary dormancy. Results: All species had long-term persistent seed banks, with estimated half-lives between 3.5 and 5.5 years. Physiological dormancy was broken by stratification prior to embryo growth, during the first year of burial. Seeds were able to germinate after retrieval, in light and at warm temperatures. Only low levels of germination occurred in situ over three years of burial, and did not increase over time. During germination trials, embryos remained underdeveloped unless seeds germinated, indicating that seeds were committed to germination once embryo growth was initiated. Conclusions: Long-lived seed banks are an important trait of plants from fire-prone communities, but cues observed to promote germination in situ are not necessarily the ones that break dormancy. A failure to distinguish between these two factors can divert attention away from mechanisms actually controlling dormancy and seed bank dynamics.

Post-fire recovery of eastern bristlebirds (Dasyornis brachypterus) is context-dependent

In late December 2003, a wildfire in the Jervis Bay region of New South Wales burned through an area that previously supported a large population of the endangered eastern bristlebird (Dasyornis brachypterus). The eastern bristlebird has been described as fire-sensitive, and fire is implicated in the decline of the species. The frequency of occurrence of bristlebirds was investigated in the second week after the fire in a range of sites varying in fire intensity. Bristlebirds were found in burned habitats but were less common in the sites that were more intensely burnt. Bristlebirds had been surveyed along transects in this area two months before this fire and were surveyed again 1, 9 and 13 months after the fire. Compared with prefire numbers, bristlebird numbers decreased in burnt areas after the fire and increased in unburnt areas. This pattern was evident for up to nine months after the fire, after which bristlebird numbers returned towards prefire levels in both burnt and unburnt vegetation. This is in contrast to some previous research on bristlebirds and fire. We suggest that bristlebirds avoided the fire by moving to unburnt areas and returned later when conditions were more suitable. We consider that the apparently slight impact of this fire on bristlebirds was due to the close proximity of unburnt habitat and other refuges. The response of bristlebirds and presumably other birds to fire is likely to be strongly context-dependent, so fire management may be able to be designed so as to be compatible with the conservation of local bristlebird populations.

Characterizing the Litter in Postfire Environments: Implications for Seedling Recruitment

Litter accumulation after fire influences the magnitude of seed predation and seed germination. How litter accumulation and patchiness influence postfire seedling recruitment is poorly known. Species with persistent seed banks have seeds available for germination in the immediate postfire period. In contrast, plants with transient seed banks must flower after fire to place seeds in the postfire habitat. In southeastern Australian sclerophyll forests, most seedling recruitment occurs within 3 yr after fire. We found that less litter had accumulated in sites <6 mo postfire than in those 2–3 yr postfire. Seedlings of species with persistent seed banks therefore encounter postfire environments with large areas covered by little or no litter. Among these species there is great variation in seed mass, including some species with very light seeds. In contrast, the seed mass of transient seed bank species is less variable, with the lightest species more than 70 times heavier than the lightest persistent seed bank species. These seeds arrive 1–3 yr postfire and encounter habitats with more litter and fewer bare patches. This pattern suggests that litter accumulation after fire has influenced the evolution of seed mass either directly, by affecting germination and seedling survival, or indirectly, by affecting seed predators.

Is the post-disturbance composition of a plant population determined by selection for outcrossed seedlings or by the composition of the seedbank?

Seedbanks are expected to buffer populations against disturbances, such as fire, that could alter the genetic composition of smaller, ephemeral adult populations. However, seedling genotypes may be influenced by the spatially heterogeneous nature of both the seedbank and the disturbance (for example, germination may vary with local disturbance) and also by selection acting on germination and post-germination performance. We used microsatellite-DNA surveys of seedlings emerging from the soil-stored seedbanks of Grevillea macleayana after wildfire to compare diversity and spatial structure in seedlings and adults, and through resampling of the seedling data set, to determine whether the resultant adult population reflected the effects of selection or random seedling mortality. The large post-fire seedling cohorts captured the full allelic diversity of the pre-fire adult population. However, we found a mismatch in the genotypic structure of adults and seedlings. Seedlings displayed larger heterozygous deficits than adults; however, over the ensuing 11 years, seedling heterozygosity eventually matched values for the pre-fire adults. Increasing heterozygosity among adults has generally been attributed to heterosis and/or reduction in Wahlund effects via self-thinning. Resampling of early post-fire seedlings to generate samples of equivalent size to survivors at 11 years showed that increases in heterozygosity must be driven by selection favouring outcrossed seed. This finding is important in an evolutionary context but also has implications for the restoration of natural or managed populations where a seedbank is a viable source of recruits.

The accumulation of genetic diversity within a canopy-stored seed bank

Many plants regenerate after fire from a canopy‐stored seed bank, in which seed are housed in fire resistant confructescences (cones) that remain on maternal plants. This strategy would be favoured if plants accumulate a sufficiently large and genetically diverse seed bank during interfire intervals. We use a 16‐year demographic study and surveys of microsatellite variation to quantify and explain the rate of accumulation of genetic diversity within the canopy seed bank of the shrub Banksia spinulosa. Flowering and fruit set were highly variable. An initial sample in 1991 of 354 reproductively mature plants generated 426 cones over 16 years, of which only 55 cones from 40 maternal plants persisted until 2005. By genotyping seed from these 55 cones we demonstrated that genetic diversity accumulated rapidly within the seed bank. Resampling revealed that diversity was determined by the number, not the age, of cones. Cones were widely distributed among plants, outcrossing rates were high (mean tm = 1.00 ± 0.04) and biparental inbreeding low. Adults displayed little evidence of isolation by distance and the genotypic diversity of seed cohorts was independent of the density of neighbouring potential sires. We therefore estimate that within at least 13 individual years the number of cones produced per year (14–63) would have contained 100% of the adult genetic diversity. We conclude that a highly outcrossed mating system and relatively widespread pollen dispersal ensure the rapid development of a genetically diverse and spatially and temporally homogeneous seed bank.

Genetic tests of the isolation of rare coastal dwarf populations of Banksia spinulosa

In southern New South Wales, a suite of widespread plant species exhibit short-statured ‘dwarf’ growth forms on coastal headlands. It is unclear whether such populations are genetically distinct or whether dwarfism is a plastic response to the environment. We used four microsatellite markers to assess genetic differentiation among populations from coastal and inland sites for Banksia spinulosa var. spinulosa. We sampled plants from six locations, including from three ‘dwarf’ and three ‘normal’ populations. Mean levels of genetic diversity were slightly higher in the forest (Na = 7.07 ± 0.25; He = 0.80 ± 0.09) than on the coast (Na = 5.92 ± 0.70; He = 0.72 ± 0.10). In general, populations displayed genotypic diversity expected for outcrossed sexual reproduction, with 161 of 172 individuals displaying unique genotypes and mean values of FIS close to zero. However, we found evidence of at least limited clonal replication in each of four populations and, within one coastal population, 11 of 27 individuals displayed one of three replicated genotypes, implying that the effective population size may be considerably smaller than would be inferred from the number of plants at this site. Relative to studies with other Proteaceae, this set of populations showed low, although significant, levels of differentiation (global FST = 0.061; P < 0.001), with extremely low, although significant, divergence of forest and coastal populations (FRT = 0.009; P < 0.001). There was no evidence of isolation by distance. These data imply that coastal dwarf populations are genetically similar to more extensive inland populations but in at least one case, may be at a greater risk of extinction because of low effective population size.

Window of opportunity: an episode of recruitment in a Banksia hybrid zone demonstrates continuing hybridization and phenotypic plasticity.

BACKGROUND AND AIMS In perennial plants (especially post-fire resprouters), extant populations may reflect recruitment events in the distant past. This is true of hybrid zones formed by two Banksia species of swamps and woodlands in south-eastern Australia, Banksia robur and B. oblongifolia. Both resprout after fire but recruitment is dependent on periodic fires. Although plants of intermediate morphology have also been identified as hybrids using allozyme markers, the extent of ongoing hybridization is unknown. This study investigates whether both microsatellite markers and morphological measurements can be used to distinguish between the two species and their hybrids. A recent recruitment event and microsatellite markers allow the frequency of ongoing hybridization to be estimated, and also the effects of environmental variation on the morphology of plants and seedlings to be tested. METHODS Variation at seven microsatellite loci was scored and seven leaf characteristics within putatively pure stands and mixed stands of both species were measured, revealing that the two species were genetically and morphologically distinct and that mixed stands also contained genetically and sometimes morphologically distinct hybrids. An opportunity created by wildfires was used to analyse the genetics and morphometrics of adults and seedlings from two hybrid zones. KEY RESULTS Approximately 9 % of adults and 21 % of seedlings were identified as genetic hybrids in both hybrid zones. Within these sites, the genotype of mature plants correlated well with morphology, except for some hybrid plants that had parental morphology. However, seedling morphology was highly variable and insufficient to describe the composition of the hybrid zone in this cohort. Greater phenotypic plasticity was evident among seedlings growing within the hybrid zones than seedlings growing in pots. CONCLUSIONS The hybrid zones are complex and the range of genotypes detected in seedlings reveals both continuing hybridization and introgression.

The coupling of recruitment and disturbance by fire in two resprouting Proteaceae species

Recruitment in plant populations is often tightly coupled to major disturbances such as fires. For species with persistent seed banks, fire-related cues may allow or enhance germination. The litter layer influences germination and may modify the impact of seed predators on seeds and seedlings. The litter layer is obviously affected by fire, providing one mechanism by which disturbance can determine recruitment. We tested the role of litter in the disturbance–recruitment coupling of two species with contrasting seed release timing after fire—Banksia serrata (canopy seed bank) and Telopea speciosissima (transient seed bank) by planting their seeds both early and late in the post-fire recruitment period (PRP) and manipulating litter density in orthogonal treatments. Vertebrate seed predators were excluded. Both species established more seedlings late in the PRP, although results were strongly influenced by very poor establishment at one site. Invertebrate seed predators consumed more T. speciosissima seeds in sites early (69.5%) than late in the PRP (51.2%), while consumption of B. serrata seeds was lower overall and comparable across sites (average 47.3%). Surprisingly, litter had very little effect on establishment and none on invertebrate seed predation, suggesting that other factors are more important. Recruitment was only loosely coupled to disturbance for the canopy seed bank species; for the transient seed bank species, the coupling was tighter but separated in time from the disturbance. Understanding both the strength and temporal aspects of the disturbance–recruitment coupling is necessary for appropriate management of plant functional diversity in fire-prone habitats.