Copeia 2009, No. 2, 320–327
Aspects of the Biology and Population Genetics of the Antarctic Nototheniid Fish Trematomus nicolai Kristen L. Kuhn1, Thomas J. Near1, Christopher D. Jones2, and Joseph T. Eastman3 Trematomus nicolai is a near shore benthic notothenioid fish most abundant in the subzero shelf waters of East Antarctica. During recent collecting we obtained the first specimens of this species from West Antarctica (the Bransfield Strait), and we compare these with specimens from the Ross Sea (McMurdo Sound) in East Antarctica. Because T. nicolai has been frequently misidentified as T. tokarevi, we provide several non-meristic characters that separate these species. We employ a radiographic technique for rapid visualization of the cephalic lateral-line canals, an important diagnostic character in trematomids. Compared to those from McMurdo Sound, the Bransfield Strait specimens have lower ranges and mean counts for meristic characters, with significant differences for anal rays, pectoral rays, and vertebrae. Our data suggest a panmictic population, but the Bransfield Strait specimens live in water 3–4uC warmer than McMurdo Sound, and this may contribute to lower meristic counts. The mean buoyancy between the two samples is not significantly different. We examined sequence variation in the ND2 portion of the mitochondrial DNA (mtDNA) genome for evidence of population structure in samples from both areas. We identified 12 mtDNA haplotypes (haplotype diversity [h] = 0.978, nucleotide diversity [p] = 0.458%) and analysis of molecular variance (AMOVA) shows no significant global differentiation. A medianjoining network that represents the genealogical relationships among the mitochondrial haplotypes also shows little separation between the samples from West and East Antarctica, and additional tests suggest the T. nicolai population is in equilibrium and of constant size. Trematomus nicolai exemplifies the potential of the Antarctic current regime for circumAntarctic dispersal of a variety of organisms in the Southern Ocean.
A
BOUT 100 species of notothenioid fishes occur in the cold waters of the Southern Ocean surrounding Antarctica (Eastman, 1993). With little competition from a sparse non-notothenioid fauna, they underwent phyletic as well as morphological and ecological diversification as Antarctic waters became isolated (Eastman, 2000). Much of this diversification occurred during the past 24 million years (Near, 2004). Notothenioids monopolize species diversity (76.6%), abundance (91.6%), and biomass (91.2%) on the Antarctic shelf, a level of dominance by a single taxonomic group that is unique among piscine shelf faunas of the world (Eastman, 2005). This diversification is also noteworthy because notothenioids lack a swim bladder yet occupy pelagic, cryopelagic, semipelagic, and benthopelagic habitats in addition to the ancestral benthic habitat (Eastman, 1993). During recent collecting in the Bransfield Strait and around the northern Antarctic Peninsula, we obtained specimens of Trematomus nicolai (Fig. 1), one of the least studied Trematomus species, from areas substantially outside of its documented geographic distribution (DeWitt et al., 1990). Study of these West Antarctic specimens, and comparison with those from East Antarctica, allowed us to fill in several aspects of the biology of T. nicolai including morphological variation and population genetic structure. The monophyly of notothenioids is well supported in phylogenetic analyses of morphological and molecular data (Balushkin, 2000; Near et al., 2004; Near and Cheng, 2008). Phylogenies inferred from both mitochondrial and nuclear gene DNA sequence data suggest that the nototheniid clade Trematomus is monophyletic only when it includes Pagothenia and Cryothenia (Ritchie et al., 1996; Cziko and Cheng, 2006; Sanchez et al., 2007; Near and Cheng, 2008). Trematomus, a clade comprising 15 species, is the character-
1
istic faunal element in the subzero waters of the East Antarctic shelf (DeWitt, 1971), and is an important component of notothenioid abundance and biomass in these areas (Ekau, 1990). Nevertheless, there are gaps in our knowledge of trematomids and, with the exception of work by Janko et al. (2007) that we will consider later, little is known about the population genetic structure of most species. In this paper we will present information on species identification, meristic variation, and taxonomic characters, including a rapid radiographic technique for examining the bony cephalic lateral-line canals, biogeography and buoyancy, and population structure of specimens sampled from West and East Antarctica estimated from mitochondrial DNA sequence data. MATERIALS AND METHODS Collection of specimens and tissue samples.—We obtained 11 specimens of T. nicolai with bottom trawls during a 2006 survey conducted by the US Antarctic Marine Living Resources (AMLR) Program aboard the RV Yuzhmorgeologiya. We collected the specimens during February and March at seven stations in the Bransfield Strait/northern Antarctic Peninsula area of West Antarctica (62.4–63.3uS, 54.9– 59.5uW; Fig. 2). Depth of capture was 169–460 m. For comparison we used 30 specimens from McMurdo Sound in the Ross Sea, East Antarctica (77u519S, 166u409E). These were captured during October and November 1974–1979 in bottom traps at 30–50 m. All specimens are deposited in the Yale University Peabody Museum of Natural History (YPM). Institutional abbreviations are as listed at http://asih.org/ codons.pdf. We also obtained muscle tissue samples from the Bransfield Strait specimens during the AMLR 2006 cruise
Department of Ecology and Evolutionary Biology and Peabody Museum of Natural History, Yale University, New Haven, Connecticut 06520-8105; E-mail: (KLK)
[email protected]; and (TJN)
[email protected]. Send reprint requests to KLK. 2 Southwest Fisheries Science Center, National Marine Fisheries Service, National Oceanic and Atmospheric Administration, 8604 La Jolla Shores Drive, La Jolla, California 92037-1508; E-mail:
[email protected]. 3 Department of Biomedical Sciences, Ohio University, Athens, Ohio 45701-2979; E-mail:
[email protected]. Submitted: 9 May 2008. Accepted: 22 December 2008. Associate Editor: J. M. Quattro. DOI: 10.1643/CG-08-087 F 2009 by the American Society of Ichthyologists and Herpetologists
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Fig. 1. Live specimen of Trematomus nicolai (YPM 16520, SL 5 209 mm) from the Bransfield Strait showing color, body conformation, and tubular scales of the middle lateral line near the caudal peduncle. The protruding premaxillary teeth are also visible.
and from two separate collections of specimens from McMurdo Sound made in different years (1998 and 2001– 2003) but at the same locality given above. Tissue samples were either frozen or stored in 95% ethanol prior to DNA extraction. Meristic analysis.—Following methods of Hubbs and Lagler (2004), we made seven fin ray, scale, and vertebral counts on each specimen from the two populations; however, we followed the standard practice for notothenioids and individually counted the last rays of the dorsal and anal fins (DeWitt, 1966). Counts were made with the aid of a dissecting microscope and radiographs. We made measurements of buoyancy on heavily anesthetized specimens in the field (Eastman and Sidell, 2002; Near et al., 2003). The buoyancy sample consisted of a subset of eight specimens from the Bransfield Strait (water temperature 5 0–2uC) and 16 from McMurdo Sound (water temperature 5 21.9uC).
Fig. 2. Map showing stations where Trematomus nicolai were collected in the Bransfield Strait and northern Antarctic Peninsula area of West Antarctica.
The McMurdo sample was previously used for buoyancy measurements (Eastman and DeVries, 1982) and vertebral counts (Eastman, 1983). Differences in density between 0uC and 5uC sea water are , 0.1%. Furthermore, errors in buoyancy measurements attributable to changes in salinity, temperature, and pressure are small, totaling only about + 0.15% of the weight of fish in air (Corner et al., 1969), and usually disregarded (Denton and Marshall, 1958). In summarizing and comparing meristic and buoyancy data for the two populations, we utilized descriptive statistics and independent t-tests. Measurements were not independent since we made nine counts and measurements on each fish. We therefore used the Bonferroni adjustment to ensure that we accepted a conservative P-level (0.05/9 5 0.0056) for significance testing. In all comparisons we used a null hypothesis of no difference between the two populations in length, meristic counts, or buoyancy. We used bivariate scatter plots and least-squares regression of lntransformed data to examine the relationship between buoyancy and both standard length and body weight. We employed one-way ANOVAs (F-tests) and t-tests to evaluate the levels of significance for exponents of the regressions. Visualization of cephalic lateral-line system.—We assessed the arrangement of the cephalic lateral-line system by injecting approximately 0.5 ml of 0.1% aqueous cresyl violet acetate into pores cannulated with PE-50 tubing (0.965 mm outer diameter). For definitive examination, we cleared and stained specimens with alizarin red S (Taylor, 1967) dissolved in 75% ethanol (Springer and Johnson, 2000). The head skin was left intact to preserve the pores and canals. We also used soft radiography—production of X-rays at low voltage with filtration through a beryllium window (Miller, 1957)—as a more rapid technique for visualizing the dorsal portion of the cephalic lateral-line system. We removed the dorsal portion of the head with a horizontal cut through the angle of the jaws and a transverse cut at the anterior margin of the first dorsal fin. We placed the dorsal surface of the head against the film cassette and produced radiographs with a Hewlett-Packard Faxitron soft X-ray
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machine. We operated the machine at 30 kVp and 3.0 mA, with an exposure time of 2.3 min. We used Kodak Industrex MX125 film (medium speed, very high contrast, high definition, very fine grain) in lead-backed cardboard cassettes. Film-to-source distance was 122 cm. DNA sequencing.—DNA was obtained from 11 T. nicolai samples from the Bransfield Strait and from three individuals from the Ross Sea. DNA was extracted using the tissue protocol specified by QIAGEN’s QIAampH DNA Mini Kit (Qiagen Inc., Valencia, CA). The mitochondrial (mtDNA) encoded NADH dehydrogenase subunit 2 gene (ND2, 1047 base pairs) was amplified using the polymerase chain reaction (PCR) in 25 ml reactions containing 1 ml 25 mM MgCl, 2.5 ml 103 CoralLoad PCR Buffer (Qiagen Inc., Valencia, CA), 0.5 ml 10 mM dNTP mix, and 0.5 ml of a 10 mM stock of each GLN and ASN primer (Kocher et al., 1995), 0.3 ml Taq DNA Polymerase (Invitrogen Co., Carlsbad, CA), and 1.5 ml DNA template. Cycling conditions for PCR consisted of an initial denaturation of 2 min at 94uC, followed by 35 cycles of denaturation at 94uC for 45 sec, annealing at 55uC for 60 sec, and extension at 72uC for 90 sec, with a final extension of 5 min at 72uC. PCR products were run on 1% agarose gels to check the success of amplification and products were subsequently purified for sequencing using AMPureH beads according to the manufacturer’s protocol (Agencourt Bioscience, Beverly, MA). Cycle sequencing reactions in both directions were run using the ABI BigDyeH Terminator v3.1 Cycle Sequencing Kit following the manufacturer’s instructions (Applied Biosystems Inc., Foster City, CA) and were generated on an ABI 3700 capillary sequencer by the W.M. Keck Foundation Biotechnology Resource Laboratory at the Yale University Medical School. DNA sequences were edited and aligned using the programs SeqManII (DNASTAR Inc., Madison, WI), Sequencher v. 4.5 (Gene Codes, Ann Arbor, MI), and BioEdit (Hall, 1999). Population structure.—Mitochondrial haplotypes were examined using analysis of molecular variance (AMOVA) as implemented in ARLEQUIN version 3.11 (Excoffier et al., 2005). AMOVA evaluates patterns of molecular variation within and among populations, taking into account the sequence divergence of molecular haplotypes as well as their frequencies. The degree of population subdivision was estimated by F-statistics (Weir and Cockerham, 1984). Nucleotide (p) and haplotype (h) diversities were calculated for mitochondrial sequence data using the program DnaSP v. 4.10.9 (Rozas et al., 2003). A median joining network was generated from mitochondrial haplotypes using the program Network v. 4.0.1.7 (http://www.fluxus-engineering.com) for each species. Network files were generated from aligned DNA sequence data using the program DNA Alignment (http://www.fluxusengineering.com). Median joining networks are commonly used to depict relationships of closely related mitochondrial haplotypes or nuclear gene alleles, for which traditional phylogenetic approaches yield multiple plausible trees (Bandelt et al., 1999). Pairwise comparisons of mitochondrial sequence data were used to generate an observed mismatch distribution. This distribution may then be compared to neutral expectations under models of constant population size or historical changes in population size (Rogers and Harpending, 1992). Mismatch distributions were calculated using
Copeia 2009, No. 2
Arlequin 3.01. Fu and Li’s D* and F* tests (1993), Fu’s FS test (1997), and Tajima’s D values (1989) were calculated using the program DnaSP 4.10.4 (Rozas et al., 2003). RESULTS Identification and taxonomic characters.—Our specimens of Trematomus nicolai (Fig. 1) were originally misidentified as T. tokarevi, and the two species have frequently been confused (Andriashev, 1978; DeWitt et al., 1990). In many trematomids meristic counts overlap to the extent that they are of little use in separating species; however, T. nicolai differs from T. tokarevi in other features including a middle lateral line with tubular scales (Fig. 1), clearly shown in the illustration of the type from Cape Adare 615 km north of McMurdo Sound (Boulenger, 1902). All our specimens from both localities have this feature. Other specimens from Cape Hallett, near Cape Adare, lack tubular scales in the middle lateral line (Miller, 1993), as do those from the Queen Mary Coast (Waite, 1916), also in East Antarctica. The later specimens are probably T. tokarevi given their small size (115 mm TL) and depth of capture (655 m). An identification as T. tokarevi is probably also valid for small (180 mm TL) specimens from deep water (470–540 m) near King George Island in the South Shetlands (Sko´ra and Neyelov, 1992). An outer row of long protruding teeth on the premaxillaries and dentaries (Figs. 1, 3) is a distinctive and underutilized diagnostic character for T. nicolai. Although the teeth are not mentioned in the original description, they are clearly evident in the accompanying figure (Boulenger, 1902:plate XV). The two species also differ in color of the iris and peritoneal pigmentation. Trematomus nicolai has a brown iris (Fig. 1) and gray peritoneum, whereas T. tokarevi has a silvery iris and black peritoneum. Furthermore, unlike any other trematomid, T. tokarevi possesses a distinctive tubular fold or lobe at the anal opening (Andriashev, 1978). The cephalic lateral-line system is important in the taxonomy of Trematomus (Jakubowski, 1970). For example, the arrangement of the bony canals and pores of the supratemporal canal is the major feature in the key couplet separating T. nicolai and T. tokarevi (DeWitt et al., 1990). In T. nicolai the left and right supratemporal canals are continuous across the dorsal surface of the head, consisting of four bony canal segments and three pores (Fig. 3), whereas in T. tokarevi left and right canals are interrupted and consist of lateral bony segments (each with one pore) connected to the temporal canals. A central portion with two bony segments and three pores completes the canal. Thus the entire supratemporal canal in T. tokarevi consists of four bony segments and five pores. We found that pores and canals are inconspicuous, even in large specimens of T. nicolai, and that cannulation through the central pore and injection of dye was necessary to see the arrangement of the canals and the location of the lateral pores. Using an Alizarin-stained specimen for comparison (Fig. 3A), we employed soft radiography as a rapid method of visualizing the canals. When the dorsal head is removed and radiographed, the confusion caused by the superimposition of the ventral bones is avoided and the bony segments of the supratemporal canal are clearly evident as are parts of the infraorbital and temporal series (Fig. 3B). Taking and developing a radiograph takes 30– 45 min versus six weeks for the alizarin procedure; this is a
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lateral-line scales are highly variable and the taxonomic value of this character lies in its presence or absence. Within limits allowed by the genotype, meristic characters are influenced by environmental conditions, especially water temperature during embryonic development (Jordan, 1891), an observation known as Jordan’s rule. The lower meristic counts at the lower latitudes of the Bransfield Strait are consistent with Jordan’s rule postulating an inverse relationship between water temperature and number of meristic elements, especially vertebrae. The waters of the Bransfield Strait are approximately 3–4uC warmer than the 21.9uC waters of McMurdo Sound. While this appears to be an example of Jordan’s rule, other factors (salinity, size swimming mode, phylogenetic relationships) may contribute to the effect and the relative influence of each individual factor is difficult to apportion (McDowall, 2008). Population level studies are essential in resolving this question, especially when the sample, such as ours, is controlled in the sense that it is drawn from a potentially panmictic population (see below).
rapid and useful technique if sufficient specimens are available for removal of the dorsal head.
Size, depth distribution, and biogeography.—The specimens of Trematomus nicolai from the Bransfield Strait are significantly longer than those from McMurdo Sound (Table 1), with a 395 mm TL (1.068 kg) specimen exceeding by 35 mm the maximum known length for the species, thus placing it among the largest trematomids (DeWitt et al., 1990). Most occurrences in the Ross Sea (Eastman and DeVries, 1982) and other East Antarctic localities (Miller, 1993) are from water less than 50 m deep, but in the Weddell Sea they occur as deep as 420 m (Ekau, 1990). Our specimens from the Bransfield Strait extend the known depth range of this species to 460 m. Like many notothenioids and some trematomids, T. nicolai is eurybathic with larger specimens found at greater depths (Ekau, 1990). Thus T. nicolai is more ecologically plastic than previously suspected and joins many other notothenioids with bathymetric ranges of 500– 700 m (Andriashev, 1965).
Meristics.—Compared with the sample from McMurdo Sound, the Bransfield Strait specimens of Trematomus nicolai have lower ranges and mean counts for meristic characters (Tables 1, 2). The differences are significant only for anal rays, pectoral rays, and vertebrae. Counts for tubular middle
Buoyancy and habitat.—Values for percentage buoyancy (%B) reflect habitat in notothenioids; values range from 0% in neutrally buoyant species to 6% in heavy benthic species, with most benthic trematomids clustering at 2.9– 3.4% (Eastman, 1993; Eastman and Sidell, 2002). This first
Fig. 3. Comparison of the dorsal skull of Trematomus nicolai showing the supratemporal canal as revealed by alizarin staining (A) and soft radiography (B) in two different specimens in lot YPM 16402, SL 5 251 mm and 290 mm, respectively. Arrows indicate canal segments of supratemporal canal; arrowheads indicate canal segment of temporal canal; circles indicate location of supratemporal pores. Some canals of the infraorbital series are also visible. The distinctive long protruding premaxillary and dentary teeth are also evident.
Table 1. Comparison of Size, Meristic, and Buoyancy Data for Trematomus nicolai from West and East Antarctica. Numbers of specimens are 11 for Bransfield Strait sample and 30 for McMurdo Sound, with the exception %B where numbers are 8 and 16, respectively. Abbreviations: SL 5 standard length; D1 5 spines in first dorsal fin; D2 5 rays in second dorsal fin; A 5 anal rays; P 5 pectoral rays; Vert 5 vertebrae; ULL 5 tubular scales in upper lateral line; MLL 5 tubular scales in middle lateral line; %B 5 percentage buoyancy (weight in water/weight in air 3 100). * denotes statistical significance at the Bonferroni adjusted P-level of , 0.0056.
Bransfield Strait range SL (mm) D1 D2 A P Vert ULL MLL %B
160–335 3–4 36–38 31–33 27–29 50–51 39–42 6–24 2.84–3.97
McMurdo Sound
mean 6 SD 226 3.9 36.9 31.6 28.0 50.7 39.9 11.6 3.37
6 6 6 6 6 6 6 6 6
52 0.3 0.7 0.7 0.6 0.5 1.0 5.5 0.36
range 97–180 3–5 36–39 31–34 28–30 51–53 38–44 2–20 2.42–3.62
mean 6 SD 137 4.0 37.4 32.4 29.1 51.5 40.7 11.7 3.13
6 6 6 6 6 6 6 6 6
22 0.4 0.7 0.6 0.6 0.6 1.7 4.6 0.33
P-level 0.00001* 0.4950 0.0785 0.0036* 0.0001* 0.0002* 0.0753 0.9405 0.1080
3 1 9 2 1 3 9 5 4
41
12 8 16 3
51
6 2 21 7 3 2 1 12
1
28
5 16 5 1 1 2 11 6 16 3 2 10 26 Brans McM
1 2
2
38 4 3
D1
5
36
37
D2
39
31
32
A
33
34
27
P
29
30
50
Vert
52
2
53
3
38
39
40
ULL
42
43
1
44
Copeia 2009, No. 2 Table 2. Frequency Distribution of Meristic Counts for Trematomus nicolai from West and East Antarctica. Numbers of specimens are 11 for Bransfield Strait (Brans) sample and 30 for McMurdo Sound (McM). Abbreviations as in Table 1.
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comparison of buoyancies from geographically separated samples of any trematomid revealed no significant difference in mean buoyancy between populations of Trematomus nicolai from the Bransfield Strait and McMurdo Sound (Table 1), and values fell within the expected range for benthic trematomids. Because buoyancy values are a measure of the relative density of the fish, and reflect the transitory influences of gut contents, nutritional condition, and reproductive state (although ripe individuals are excluded from the measurements), dispersion of the values is expected. Attempting to control for the influence of these variables is not biologically realistic as they reflect normal aspects of the daily and yearly life cycles. Nevertheless, the standard deviations of the majority of the adequately sampled species are relatively small 6 0.3–0.6% (Eastman, 1993; Eastman and Sidell, 2002), and this also applies to our samples of T. nicolai (Table 1). The approximate 200–400 m difference in depth preference between the two populations is not reflected in significant differences in buoyancy, and small differences we did find probably have little functional consequence in a species without a swim bladder. When the samples from the Bransfield Strait and McMurdo Sound are pooled and %B values are plotted against either body weight or standard length, there is a slight trend for larger specimens to be more buoyant (to have a lower %B). However, least-squares regressions of %B against both weight and standard length, respectively, are not significant (r2 5 0.04, F1,22 5 0.88, P 5 0.36; r2 5 0.08, F1,22 5 1.85, P 5 0.19). A size (weight) effect on buoyancy (lower %B with increasing weight) has been documented in large samples of some nototheniids and channichthyids (Eastman and Sidell, 2002), but the most striking example is seen in the large, neutrally buoyant nototheniid Dissostichus mawsoni. As lipid accumulates with growth in this species, %B falls from 4.5% in juveniles to 0% in $ 81 cm SL adults (Near et al., 2003). Population genetics.—We examined the ND2 portion of the mtDNA genome for evidence of population structure in 14 individual Trematomus nicolai samples collected from the Bransfield Strait in West Antarctica and the Ross Sea in East Antarctica (11 from the Bransfield Strait and three from the Ross Sea). Twelve mtDNA haplotypes were identified (haplotype diversity [h] 5 0.978, nucleotide diversity [p] 5 0.458%). In total, 28 variable nucleotide positions were identified, with nine of the polymorphisms being nonsynonymous with regard to inferred amino acid sequences and all were observed in single individuals. AMOVA showed no significant global differentiation (FST 5 0.000, P 5 0.486) for the full data set. Additionally, a median-joining network illustrating genealogical relationships among the mitochondrial haplotypes shows little separation between West and East Antarctica localities (Fig. 4). Neither Fu and Li’s D* and F* tests, nor Fu’s FS and Tajima’s D values, were significant for the pooled collections of T. nicolai (22.123, 22.213, 22.022, and 21.677, respectively). The mismatch distribution of mitochondrial haplotypes observed resembles the ragged, multimodal pattern (data not shown) expected from an equilibrium population of constant size (Rogers and Harpending, 1992; Kuhn and Gaffney, 2006). DISCUSSION Biology of Trematomus nicolai.—Samples of Trematomus nicolai from the Bransfield Strait represent a major range
Kuhn et al.—Biology of Trematomus nicolai
Fig. 4. Median-joining network for Trematomus nicolai ND2 haplotype data. Circle sizes are proportional to the number of individuals possessing the haplotype. Each color represents a different haplotype locality (black represents the Bransfield Strait, gray represents the Ross Sea). Small unfilled circles represent unobserved haplotypes. Hatch marks represent mutational steps separating haplotypes.
extension from primarily high latitude shelf localities in East Antarctica (DeWitt et al., 1990) to the northern Antarctic Peninsula. While there are putative specimens from Petermann Island (Roule and Despax, 1911) and King George Island in the South Shetlands (Sko´ra and Neyelov, 1992), and from the South Orkney Islands (Matallanas, 1997), these are questionable given past confusion over identification and the absence of cited museum catalog numbers for these specimens. Like all the high Antarctic notothenioids, Trematomus nicolai has body fluids fortified by antifreeze glycoproteins for life at subzero temperatures (DeVries and Cheng, 2005). Despite living at 21.9uC in McMurdo Sound, one of the most thermally stable marine environments, several species of Trematomus (sensu lato) have the capacity to rapidly acclimate to 4uC (Podrabsky and Somero, 2006; Robinson and Davison, 2008). Thus, the range of habitat temperatures encompassing populations of T. nicolai is well within tolerances for the genus. There is a near shore/off shore dichotomy in the bathymetric ranges of trematomids (Andriashev, 1965; DeWitt, 1971). Trematomus nicolai exemplifies a near-shore species (0–700 m), along with T. newnesi, T. bernacchii, and T. pennellii, because the lower end of their depth range is in extremely shallow water. The off shore trematomids found at 70–1,200 m (T. vicarius, T. hansoni, T. loennbergii, T. scotti, T. lepidorhinus, T. eulepidotus, and T. tokarevi), while sometimes found near shore, usually occur in deeper waters near the outer shelf and upper slope. While both groups consist of benthic species with mid-range buoyancy values, each also includes a more buoyant species. For example, T. newnesi (%B 5 2.62) is a semipelagic near shore species and T. loennbergii (%B 5 2.28) is an epibenthic off shore species. These two species are not closely related to other Trematomus species, reflecting independent evolution of water column species in Trematomus and possibly other major lineages in the Nototheniidae (Sanchez et al., 2007; Near and Cheng, 2008). With regard to ecology, diversification in buoyancy is the hallmark of the nototheniid radiation.
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Population genetics and dispersal.—Previous population genetics work by Janko et al. (2007) evaluated sequence data from four Trematomus species, with different life-history strategies, from the high Antarctic. Despite small sample sizes, they identified a pattern of mitochondrial haplotype structure in populations of T. bernacchii sampled in the South Shetland Islands (WST 5 0.699, P , 0.05) versus those sampled from the Ross Sea; however, they found no genetic structuring among these two populations using the nuclear intron ribosomal protein S7 (Janko et al., 2007). There was no mitochondrial or nuclear gene population structure in sampled populations of T. pennellii, T. newnesi, and T. borchgrevinki (Janko et al., 2007). This is consistent with the results obtained here suggestive of no global population structure in T. nicolai (FST 5 0.000, P 5 0.486) for the full data set, despite the small sample size (n 5 14). The mismatch distribution observed in T. nicolai from mitochondrial sequence data is suggestive of a population of stable size. The Janko et al. (2007) examination of four near shore species found a similar pattern of constant growth and stable population size in T. newnesi, while they found evidence for expansion events or selective sweeps during the Pleistocene in T. bernacchii, T. pennellii, and T. borchgrevinki. Although we sampled a small number of individuals from two widely separated populations, our results clearly show that fixed differences were not detected and thus strong population structure was not present. Our data also suggest that populations of near shore benthic T. nicolai may be more dynamic, and the larvae more vagile, than previously expected. Our morphological and genetic data from Bransfield Strait suggests a circum-Antarctic distribution in Trematomus nicolai. Thus eight of the traditionally recognized 11 species of Trematomus now have documented circum-Antarctic distributions. The Antarctic continent is encircled by the eastward flowing Antarctic Circumpolar Current (Orsi et al., 1995) and the Ross and Weddell seas also contain large gyres. There is considerable potential for advection of notothenioid larvae by this extensive current system. Marine species with planktonic larval, juvenile, or adult stages have the potential for dispersal over long distances leading to geographic homogeneity of neutral genetic markers; indeed, marine species tend to show less geographic differentiation than their anadromous or freshwater counterparts (Ward et al., 1994; Kuhn and Gaffney, 2006). Trematomid larvae have pelagic stages lasting several months to a year and thus have the potential for wide dispersal by currents (Loeb et al., 1993). On the other hand, notothenioids with fidelity to home ranges or spawning sites at isolated island groups at the periphery of the Southern Ocean, as in the case of the Patagonian toothfish Dissostichus eleginoides, tend to show more population subdivision (Rogers et al., 2006; Kuhn, 2007). Nevertheless the importance of oceanic dispersal has been underestimated (de Queiroz, 2005) and now, with rapid DNA sequencing methodology and growing databases, there are well-documented examples of dispersal for a variety of organisms in the Southern Ocean (Greve et al., 2005; Sanmartı´n et al., 2007; Waters, 2008). MATERIAL EXAMINED Trematomus nicolai. Museum specimens. Bransfield Strait/ northern Antarctic Peninsula. YPM 16393, 16402 (n 5 5), 16475, 16512, 16520, 16539, 16562; Ross Sea, McMurdo
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Sound. YPM 18359 (n 5 7), 18360 (n 5 5), 18361 (n 5 14), 18362 (n 5 4). Frozen tissue. Tissue samples are deposited in the Yale Fish Tissue Collection (YFTC) and cross referenced (in parentheses) to a specimen in the YPM collection. Bransfield Strait/northern Antarctic Peninsula. YFTC 7735 (YPM 16402), YFTC 7740 (YPM 16402), YFTC 7750 (YPM 16402), YFTC 7751 (YPM 16402), YFTC 7752 (YPM 16402), YFTC 7771 (YPM 16520), YFTC 7816 (YPM 16393), YFTC 7879 (YPM 16562), YFTC 7968 (YPM 16539), YFTC 7982 (YPM 16475), YFTC 7997 (YPM 16515). Ross Sea, McMurdo Sound. YFTC 1396, 12068, 12069 (no voucher specimens). Trematomus tokarevi. Museum specimens. Northern South Shetland Islands. YPM 16639 (n 5 3). Addendum.—Trawling during the US AMLR cruise in February–March 2009 yielded another new locality record for Trematomus nicolai from waters around the South Orkney Islands. ACKNOWLEDGMENTS We are grateful to J. Sattler for photographing Figure 3A and to D. Pratt for assembling Figure 3. Research was conducted under protocol L01-14 as approved by the Institutional Animal Care and Use Committee of Ohio University. In collecting specimens, we adhered to provisions of the Antarctic Conservation Act. For transshipping and importing specimens, we had permits from the New Zealand Ministry of Agriculture and Fisheries (#A.S. 7015 and A.S. 7016) and the United States Fish and Wildlife Service (a cleared Form 3-177). Supported by NSF grant ANT 04-36190 (JTE). LITERATURE CITED Andriashev, A. P. 1965. A general review of the Antarctic fish fauna, p. 491–550. In: Biogeography and Ecology in Antarctica, Monographiae Biologicae, Vol XV. P. van Oye and J. van Mieghem (eds.). Junk, The Hague. Andriashev, A. P. 1978. Trematomus tokarevi, a new species of the family Nototheniidae (Pisces) from the abyssal waters near Antarctica. Journal of Ichthyology 18:521–526. Balushkin, A. V. 2000. Morphology, classification, and evolution of notothenioid fishes of the Southern Ocean (Notothenioidei, Perciformes). Journal of Ichthyology 40, Supplement 1:S74–S109. Bandelt, H-J., P. Forster, and A. Roehl. 1999. Medianjoining networks for inferring intraspecific phylogenies. Molecular Biology and Evolution 16:37–48. Boulenger, G. A. 1902. Pisces, p. 174–189. In: Report on the Collections of Natural History Made in the Antarctic Regions During the Voyage of the ‘‘Southern Cross’’. British Museum (Natural History), London. Corner, E. D. S., E. J. Denton, and G. R. Forster. 1969. On the buoyancy of some deep-sea sharks. Proceedings of the Royal Society of London 171B:415–429. Cziko, P. A., and C.-H. C. Cheng. 2006. A new species of nototheniid (Perciformes: Notothenioidei) fish from McMurdo Sound, Antarctica. Copeia 2006:752–759. de Queiroz, A. 2005. The resurrection of oceanic dispersal in historical biogeography. Trends in Ecology and Evolution 20:68–73. Denton, E. J., and N. B. Marshall. 1958. The buoyancy of bathypelagic fishes without a gas-filled swimbladder. Journal of the Marine Biological Association of the United Kingdom 37:753–767.
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