Genetic diversity and population genetic structure in fragmented Allocasuarina verticillata (Allocasuarinaceae) – implications for restoration

Vegetation restoration in fragmented regions is constrained by limited supplies of high quality seed and an understanding of the scale over which seed can be moved without causing negative outcomes. ‘Local’ seed is often prescribed for restoration but in fragmented landscapes this restricts collecting to small, inbred populations. Six polymorphic microsatellites were used to examine genetic diversity and population genetic structure in seed collected from 18 fragmented natural populations and three restored populations of the wind-pollinated and dispersed tree Allocasuarina verticillata, a key restoration species. Smaller populations produced seed crops with significantly fewer alleles, lower allelic richness and less gene diversity. Most of the populations assessed, including the restored sites, produce genetically diverse seed crops suitable for restoration but smaller populations (<30 plants) should be augmented with seed from larger populations. Principal coordinate analysis, graph-theory and Bayesian analyses found little evidence of spatially predictable genetic structure across the study region, which probably reflects long distance gene dispersal preventing the development of strong spatial structure. The absence of strong spatial patterns suggests that seed can be moved beyond current 5–50-km limits while being mindful of strong selection gradients or conditions that might indicate locally adapted populations.

Post-Transcriptional Gene Silencing in the Root System of the Actinorhizal Tree Allocasuarina verticillata

In recent years, RNA interference has been exploited as a tool for investigating gene function in plants. We tested the potential of double-stranded RNA interference technology for silencing a transgene in the actinorhizal tree Allocasuarina verticillata. The approach was undertaken using stably transformed shoots expressing the β-glucuronidase (GUS) gene under the control of the constitutive promoter 35S; the shoots were further transformed with the Agrobacterium rhizogenes A4RS containing hairpin RNA (hpRNA) directed toward the GUS gene, and driven by the 35S promoter. The silencing and control vectors contained the reporter gene of the green fluorescent protein (GFP), thus allowing a screening of GUS-silenced composite plantlets for autofluorescence. With this rapid procedure, histochemical data established that the reporter gene was strongly silenced in both fluorescent roots and actinorhizal nodules. Fluorometric data further established that the level of GUS silencing was usually greater than 90% in the hairy roots containing the hairpin GUS sequences. We found that the silencing process of the reporter gene did not spread to the aerial part of the composite A. verticillata plants. Real-time quantitative polymerase chain reaction showed that GUS mRNAs were substantially reduced in roots and, thereby, confirmed the knock-down of the GUS transgene in the GFP+ hairy roots. The approach described here will provide a versatile tool for the rapid assessment of symbiotically related host genes in actinorhizal plants of the Casuarinaceae family.

Casuarina ecology: factors limiting cone production in the drooping sheoak, Allocasuarina verticillata

Allocasuarina verticillata is an important species for biodiversity conservation on Kangaroo Island (South Australia) because it is the primary food source for the endangered glossy black-cockatoo, Calyptorhynchus lathami halmaturinus. Two potentially limiting factors, pollen and soil nutrients, were studied in the context of A. verticillata as foraging habitat for glossy black-cockatoos. Cone production was not limited by the amount of pollen available to female plants. The soils on which A. verticillata occurs on Kangaroo Island were low in nutrients. Available N, P and K were significantly increased via the application of slow-release fertiliser and the added nutrients resulted in a corresponding increase in the productivity of A. verticillata. The additional nutrients increased the number of cones produced on female branches, branch growth and potentially therefore, tree size. Since cone profitability appears to increase with tree size, the additional growth may also result in greater cone profitability in the long term. Adding slow-release fertiliser to small female A. verticillata trees and revegetation on sites with higher concentrations of soil nutrients would benefit the cockatoos. This is because other studies have shown that the cockatoos increase their foraging efficiency by cropping cones from large trees with greater cone profitability and branches with high densities of cones.

The glossy black-cockatoo (Calyptorhynchus lathami halmaturinus) spends little time and energy foraging on Kangaroo Island, South Australia

The endangered Kangaroo Island glossy black-cockatoo (Calyptorhynchus lathami halmaturinus) relies entirely on the seeds of the drooping sheoak (Allocasuarina verticillata) for food. The time budget of the glossy black-cockatoos and their foraging behaviour was recorded to provide an indication of whether their food supply was likely to be limiting. The foraging behaviour of non-breeding and breeding cockatoos was also compared to record the strategy they used to collect the additional energy needed to raise young. Glossy black-cockatoos spent a relatively small proportion of their time foraging, suggesting that the food supply was abundant in the habitats used for feeding. Non-breeding birds spent only 26% of their time feeding and breeding birds spent only 36% of their time feeding. The cockatoos spent 0.4% of their time flying, foraged in a mean of only five trees per day and harvested cones in no more than five bouts per tree. This shows that the cockatoos made few movements between drooping sheoaks and within the canopy of the sheoaks when foraging. When breeding, the cockatoos spent significantly more time per day foraging, cropped cones in significantly more bouts per tree and harvested significantly more cones per tree than non-breeding birds. This shows that breeding birds increased their energy intake without greatly increasing movement between trees. The small number of movements made by glossy black-cockatoos when foraging on Kangaroo Island reflects the abundance of food trees and may be a strategy to reduce the risk of predation.