Leucothoe eltoni sp. n., a new species of commensal leucothoid amphipod from coral reefs in Raja Ampat, Indonesia (Crustacea, Amphipoda)

Abstract A new species of leucothoid amphipod, Leucothoe eltoni sp. n., is described from coral reefs in Raja Ampat, Indonesia where it inhabits the branchial chambers of solitary tunicates. With an inflated first gnathopod superficially resembling the genus Paraleucothoe, this new species has a two-articulate maxilla 1 palp characteristic of the genus Leucothoe. While described from coral reef environments in tropical Indonesia and the Philippines, it is an established invasive species in the Hawaiian Islands. The most likely mode of introduction was a US Navy dry dock transported to Pearl Harbor in 1992 from Subic Bay, Philippines.


Introduction
While leucothoid amphipods are frequently encountered in marine faunal surveys and inventories information about their invertebrate hosts is rarely known and infrequently documented. The Leucothoidae, once thought to be a cosmopolitan and widespread taxon are now known to be a highly habitat-specific species complex. Taxonomic clarity within the group has been hampered by widespread and incorrect records of Leucothoe spinicarpa (Abildgaard, 1789). Upon further analysis many of these records are now proving to be discrete species, thus diminishing the cosmopolitan concept within the group (Krapp-Schickell and De Broyer 2014).
Because leucothoids lack a dispersive larval stage and frequently inhabit internal chambers of sessile invertebrate hosts they are potentially informative proxies for evolutionary diversity. Recent developments within the taxonomy of the Leucothoidae include: (1) availability of an electronic taxonomic database (Thomas and White 2014); and (2) widely deployed specialized in-situ underwater collecting methodologies. Specialized collection methods that isolate hosts and commensals together (sponges, ascidians, and bivalve mollusks) has led to re-examination of existing collections and spurred new efforts resulting in an increase in recently described leucothoid taxa for study and research (Myers 2013, Krapp-Schickel and DeBroyer 2014, Ortiz and Winfiled 2012, Thomas 1979, 1997a,b, Thomas and Klebba 2006, 2007, Thomas and Krapp-Schickel 2011, Thomas and Taylor 1981, White and Thomas 2009, White and Reimer 2012a, 2012b, 2012c, Winfield and Alvarez 2009 including new approaches for molecular analysis (White 2010, 2011a,b, White and Reimer 2012d. With their distinct morphology and common occurrence in shallow coastal marine environments, leucothoid amphipods drew the attention of early naturalists, resulting in some of the earliest recorded amphipod descriptions. While important taxonomic records, these early descriptions were often inadequately illustrated and described contributing to subsequent taxonomic confusion in the group. With increasing concerns about global climate change and loss of marine biodiversity, leucothoid amphipods are sensitive model organisms highly susceptible to a variety of toxicants and pollutants (Reish and Barnard 1969) and capable of providing a comparative diversity framework and serving as measures of change in marine ecosystems (Thomas 1993(Thomas , 1997b. While the lack a larval stage limits widespread dispersal, some tube-dwelling and fouling community amphipods attain broad distributions as drifters on seaweed and algae, attached to floating debris, as components of fouling communities, and their occurrence in ballast water (Carlton 2010). Leucothoids, with their constrained distributions and commensal life history, can serve as valid indicators of environmental change and sentinels of lineage-based evolutionary history (Thomas 1997a). The use of leucothoids in such context depends on precise taxonomies and representative collections across broad marine habitats. This increased precision in determining composition and assessing threat levels is of interest especially for increasingly impacted coral reef systems (Thomas 1993(Thomas , 1997b. Assessments incorporating cryptic biota could provide more refined and detailed insights than traditional coral reef inventories that incorporate organisms with large-scale dispersal capabilities (Hoeksema 2007).
Currently the revised Leucothoidae (sensu stricto) comprises 176 species in five genera. This includes 42 former anamixid species in Anamixis Stebbing, 1897 (23 spp.); Nepanamixis Thomas, 1997 (4 spp.); and Paranamixis Schellenberg, 1938 (15 spp.); and 134 leucothoid species comprised of Leucothoe Leach (132 spp.), and Paraleucothoe Stebbing, 1899 (2 spp.). Species in the former anamixid genera differ from other leucothoids in exhibiting radical sexual dimorphism, eusocial and harem guarding population structure, and tropic to warm temperate distributions. Species in Leucothoe and Paraleucothoe exhibit minor to moderate sexual dimorphism, and have tropic to polar distributions. Recent 18S rDNA sequence data by White and Reimer (2012d) suggest that the generic boundaries and definition of Paraleucothoe should be evaluated in the light of new molecular and morphological data.

Materials and methods
Using SCUBA and specialized underwater collecting techniques amphipods were sampled in-situ from ascidians, sponges, and bivalves throughout Raja Ampat, Indonesia. Specimens were captured in-situ directly from their host either with a modified squirt bottle or by isolating hosts and substrata underwater in plastic bags and later coercing the amphipods from the host using a small amount of freshwater or formalin in the lab.
Specimens were either fixed in 2% buffered formalin or 70% ethanol. Prior to observation, specimens were gently cleaned with small sable hair brushes, and transferred to glycerin for dissection, illustration, and analysis. For SEM analysis, specimens were rehydrated to distilled water (three fluid changes for 10 minutes each), soaked in a dilute surfactant for 15 minutes (two drops of Tween 80 in 100 ml of water), briefly sonicated (10 seconds) to remove accumulated surface debris, and re-rinsed in distilled water (three fluid changes for 10 minutes each). This preparation protocol was modified from Felgenhauer (1987) by using a more finely graded alcohol series (5%, 10%, 15%, 25%, 35%, 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, and 100%) to prevent distortion and shrinkage. Specimens were then fixed in salt water buffered osmium tetroxide (equal parts, under fume hood) for 2.5 hours, dehydrated in a graded alcohol series, transferred to acetone (three fluid changes for 10 minutes each), soaked in Hexamethyldisilazane Reagent (HMDS) for 15 minutes, air-dried overnight, and sputter coated with palladium for scanning electron microscopy. Photographs were taken with an ISI-DS-130 dual state scanning electron microscope. Capital letters to the right of each caption refer to the following: L = left, R = right. Lower case letters to the left of capital letters refer to the following adjectives: l = lateral, m = medial, x = magnified. Numbers to the right of capital letters refer to specific structures. "LW" in text refers to length/width ratios. Sexes are indicated by ♂ and ♀ symbols.

Figure Legend
Material is deposited at the National Museum of Natural History, Leiden (RMNH) and at the Zoological Museum of Bogor (MZB) Indonesian Institute of  Thomas, collector. Etymology. The specific epithet, eltoni, in honor of the rock musician Sir Elton John. Specifically, in reference to the large shoe-like first gnathopod of this species and the oversize boots Elton John wore as the local pinball champion in the movie "Tommy" (1975).
Description of female paratype B, 7.35 mm. Similar to males except for gnathopods 1 and 2. Gnathopod 1, carpus and propodus not greatly inflated, propodus slightly swollen distally. Gnathopod 2, palm of propodus lacking distinct tuberculation and concavities found in males.  Relationships. Leucothoe eltoni sp. n. most closely resembles L. tumida of Myers (2013) in the inflated carpus and propodus of gnathopod 1; in the short stubby antennae; pereopods 5-7 with article 4 extending more than 50 percent along posterior margin of article 5; and a 2-segmented maxilliped palp. L. tumida differs from L. eltoni in having a large excavation in the palm of gnathopod 2; and in having a smooth inner margin of the maxilliped outer plate. Both species differ in host preferences with L. tumida found in the mantle cavity of the winged pearl oyster Pteria penguin while L. eltoni prefers branchial chambers of large solitary ascidians, especially Herdmania and Polycarpa species.
Both L. tumida and L. eltoni superficially resemble members of the genus Paraleucothoe in the large inflated gnathopod 1 of terminal males. However, Paraleucothoe differs from all species of Leucothoe in having the outer plate of the maxilliped extended distally beyond palp article 1. Paraleucothoe novaehollandiae (Haswell, 1879) also has a uniarticulate maxilla 1 palp but this feature is no longer unique to the genus as a number of recently described Leucothoe species have this feature. Paraleucothoe novaehollandiae is reported from the branchial chambers of the stalked tunicates Pyura spinifera and P. praeputialis (formerly P. stolonifera) in southern Australia waters (Lowry et al 2000) and other large solitary tunicates such as Herdmania sp. The exact placement of Paraleucothoe flindersi described by Stebbing (1888) from the Torres Straits remains problematic as it lacks the extended apical lobe of maxilliped outer plate typical of L. novaehollandiae, but has a uniarticulate palp and gnathopod 1 reminiscent of L. eltoni females and juvenile males. Further resolution awaits examination of material from the type locality.
Ecology. Coral reefs, coral rubble, found primarily in branchial baskets of solitary tunicates such as Herdmania and Polycarpa sp., rarely in bivalve mollusks (winged pearl oyster Pteria penguin), and branched yellow rope sponges Callyspongia (species undetermined).
Discussion. While the native range of L. eltoni sp. n. encompasses shallow coral reef habitats in Indonesian and the Philippines, it is also an established invasive in Hawaiian waters (Coles et al. 1999). The most likely vector for introduction was a dry dock, USS Machinist, transported to Pearl Harbor from Subic Bay, Philippines in 1992. Prior extensive treatment of Hawaiian amphipods by J.L. Barnard (1970Barnard ( , 1971 and ongoing monitoring by the Bishop Museum did not document any leucothoid resembling L. eltoni sp. n. prior to 1992. Ongoing sampling of marine flora and fauna by the Bishop Museum first reported this species in 1997 as Paraleucothoe flindersi. Muir (1997) speculated it was most likely an introduced species. Such rafting on floating metal objects is a possible means of transportation for benthic marine organism (Cairns 2000;Creed andde Paula 2007, Hoeksema et al. 2012). Since first reported from Pearl Harbor in 1997 (as Paraleucothoe flindersi) L. eltoni sp. n. has spread throughout Oahu and other islands including Molokai. The author has collected L. eltoni sp. n. from sponges in Kaneohe Bay, Ohau. The effects, if any, of this species on endemic leucothoid commensals and its spread in Hawaii is unknown at this time.