Genetic Population Structure of the Hawaiian Alien Invasive Seaweed Acanthophora spicifera ( Rhodophyta ) as Revealed by DNA Sequencing and ISSR Analyses 1

Acanthophora spicifera (Vahl) Børgesen is the most widespread and invasive alien macroalga on coral reefs throughout the main Hawaiian Islands. This alga disperses from harbors and ports to coral reefs throughout the state, producing high quantities of biomass that affect a wide range of reef flora and fauna. Population samples of A. spicifera from across the main Hawaiian Islands were collected and compared through two kinds of analyses: DNA sequencing ( based on a variable region of the nuclear large subunit ribosomal RNA gene, and the mitochondrial cox 2-3 spacer region) and fragment techniques (InterSimple Sequence Repeats [ISSRs]). DNA sequencing revealed no variation for the two markers, even when collections from other areas of the Pacific and Australia were included. In contrast, ISSR analyses revealed highly structured Hawaiian populations of A. spicifera with a substantial range of both withinand among-population variation, with individual plants forming discrete clusters corresponding to geographic locality. The invasion of coral reef ecosystems by nonnative species can alter ecosystem structure and reduce indigenous biodiversity (Pandolfi et al. 2005). In Hawai‘i, the number of documented alien algal species has increased from 18 in 1992 (Russell and Balazs 1994) to 24 species in 2003 (Godwin 2003). Of the five notably invasive macroalgae, Acanthophora spicifera (Vahl) Børgesen is the most common species throughout the main Hawaiian Islands (Smith et al. 2002). Successful invaders such as A. spicifera have substantial impacts on Hawai‘i’s coral reef ecosystems (Godwin 2003). In addition to damaging native ecosystems, invasive algal blooms have caused economic losses to the state exceeding 20 million dollars (Smith et al. 2004). The widespread distribution and rapid growth of A. spicifera make it a particularly dangerous threat to Hawai‘i’s coral reefs and a priority species for control efforts. The vegetative characteristics of A. spicifera contribute to it being a successful colonizer that is capable of attaching to, and competing with, native algal species (Russell and Balazs 1994). The spiny thalli of this brittle alga are easily fragmented by wave action (Kilar and McLachlan 1986) and readily snag on suitable substrates, including other macroalgae. Kilar and McLachlan (1986) found that plants growing on exposed reef in Panama were able to generate up to 74 kg of vegetative fragments per month, and each fragment had the potential to reattach after only 2 days. Although the reproductive phenology of alien algae in Hawaiian waters has not been thoroughly investigated, A. spicifera is the only species in this category that has been consistently observed in a sexually reproductive state (Smith et al. 2002). Genetic recombination may result in increased population-level variation that allows A. spicifera to be more resistant to control efforts than clonal invasive species. Reproductive species may pose an additional threat through the release of copious quantities of microscopic spores that Pacific Science (2007), vol. 61, no. 2:223–233 : 2007 by University of Hawai‘i Press All rights reserved 1 This research was funded by the Hawai‘i Division of Aquatic Resources and the University of Hawai‘i. Manuscript accepted 12 June 2006. 2 Department of Botany, 3190 Maile Way, University of Hawai‘i at Mānoa, Honolulu, Hawai‘i 96822 (e-mail: asherwoo@hawaii.edu). 3 Corresponding author. disperse over great distances. As with many invasive organisms, the physical and reproductive characteristics of A. spicifera foster widespread dispersal and enable successful competition for resources within native reef communities. Algal invasions are often related to anthropogenic factors, including aquaculture (Abbott 1999), eutrophication (Stimson et al. 2001), hull fouling (Smith et al. 2002), ballast water discharge (Godwin 2003), and overfishing (McClanahan et al. 2001, Pandolfi et al. 2005). The major phase shifts involving primary producers on coral reefs associated with these factors have substantial and often catastrophic impacts on macroalgal, coral, and fish assemblages (Littler and Littler 1985, Hughes 1994, Russell and Balazs 1994, Stimson et al. 2001, Pandolfi et al. 2005). Acanthophora spicifera is implicated in major phase shifts on Hawaiian reefs because it has been found to produce high biomass throughout the main Hawaiian Islands (Russell and Balazs 1994). In Hawai‘i, alien algae have been documented to compete with native species of algae (Russell and Balazs 1994) and corals (Stimson et al. 2001). However, the ecological interactions involving A. spicifera on Hawaiian reefs may be complex. For example, the regular consumption of A. spicifera by Hawaiian sea turtles and herbivorous fish has been documented by Russell and Balazs (1994). Although the effects of alien algae ingestion on the diets of native herbivores are unknown (Smith et al. 2004), a large increase in edible algal biomass at a particular locale would be expected to have numerous impacts throughout the reef community. Although research concerning the effects of invasive macroalgae on tropical reefs is still limited (e.g., Coles and Eldredge 2002, Smith et al. 2004), even fewer studies have examined the genetic structure of invasive algae in tropical waters. In temperate latitudes, DNA sequencing and fragment techniques have been used to investigate invasions of green and red seaweeds. These molecular tools have been used to determine the possible geographic sources and the genetic diversity of notable invasive algae such as Caulerpa taxifolia, Grateloupia doryphora, and Asparagopsis taxiformis (Famà et al. 2002, Marston and VillalardBohnsack 2002, Chualáin et al. 2004). In the study reported here, DNA sequence analyses of a region of the nuclear ribosomal large subunit gene and the mitochondrial cox 2-3 spacer region, and Inter-Simple Sequence Repeats (ISSRs) were used to investigate the population structure of A. spicifera throughout the main Hawaiian Islands. ISSRs are a cost-effective method for the resolution of intraspecific genetic variation and have been used to study a wide variety of plant taxa (Li and Ge 2001, Sica et al. 2005), as well as other red algae (Vis 1999, Sherwood et al. 2002). Here we illustrate the utility of ISSR analyses for revealing the degree of genetic structure within and between populations, and use these data to provide insight and recommendations for management of this alien invasive seaweed. materials and methods