N 2 -fixing proteobacteria (␣ and ␥) and unicellular cyanobacteria are common in both the tropical North Atlantic and Pacific oceans. In near-surface waters proteobacterial nifH transcripts were present during both night and day while unicellular cyanobacterial nifH transcripts were present during the nighttime only, suggesting separation of N 2 fixation and photosynthesis by unicellular cyanobacteria. Phylogenetic relationships among unicellular cyanobacteria from both oceans were determined after sequencing of a conserved region of 16S ribosomal DNA (rDNA) of cyanobacteria, and results showed that they clustered together, regardless of the ocean of origin. However, sequencing of nifH transcripts of unicellular cyanobacteria from both oceans showed that they clustered separately. This suggests that unicellular cyanobacteria from the tropical North Atlantic and subtropical North Pacific share a common ancestry (16S rDNA) and that potential unicellular N 2 fixers have diverged (nifH). N 2 fixation rates for unicellular bacterioplankton (including small cyanobacteria) from both oceans were determined in situ according to the acetylene reduction and 15 N 2 protocols. The results showed that rates of fixation by bacterioplankton can be almost as high as those of fixation by the colonial N 2 -fixing marine cyanobacteria Trichodesmium spp. in the tropical North Atlantic but that rates are much lower in the subtropical North Pacific.Growth rates of plankton in open ocean surface waters are often limited by the availability of reduced forms of N. New combined N enters surface waters either by advection, diffusion of NO 3 from deep water, or biological N 2 fixation (15). The last pathway can be significant in tropical and subtropical seas, where large cyanobacteria, Trichodesmium spp., have been considered the major organisms responsible (8,9,11). Ongoing research to identify other sources of N 2 fixation and alternate pathways of reduced N to the trophic chain in the vast oceanic basins has pointed out the potential role of certain N 2 -fixing unicellular bacterioplankton, which could have a significant impact on global biogeochemistry of N and C (14, 16, 44).Previous research has identified the presence of fragments of the unicellular cyanobacterial and proteobacterial nifH genes (which encode one of the peptide molecules that form the dinitrogenase reductase subunit of nitrogenase, the enzyme responsible for N 2 fixation [31]) in the subtropical North Pacific and tropical North Atlantic oceans (16,43,44). Preliminary data on N 2 fixation rates for the 10-to 0.2-m-size fraction of the bacterioplankton have suggested that they could make an important contribution to the global N cycle (14, 44). These studies used 15 N 2 to measure 24-h N 2 fixation rates in 10-m-pore-size-prefiltered water from the subtropical North Pacific. Also, N 2 fixation rates were estimated based on unicellular phytoplankton cell numbers that came from literature values (6). Cyanobacteria are autotrophs and generate ATP necessary for N 2 fixatio...

Abstract. Coalescence theory predicts when genetic drift at nuclear loci will result in fixation of sequence differences to produce monophyletic gene trees. However, the theory is difficult to apply to particular taxa because it hinges on genetically effective population size, which is generally unknown. Neutral theory also predicts that evolution of monophyly will be four times slower in nuclear than in mitochondrial genes primarily because genetic drift is slower at nuclear loci. Variation in mitochondrial DNA (mtDNA) within and between species has been studied extensively, but can these mtDNA data be used to predict coalescence in nuclear loci? Comparison of neutral theories of coalescence of mitochondrial and nuclear loci suggests a simple rule of thumb. The ''three-times rule'' states that, on average, most nuclear loci will be monophyletic when the branch length leading to the mtDNA sequences of a species is three times longer than the average mtDNA sequence diversity observed within that species.A test using mitochondrial and nuclear intron data from seven species of whales and dolphins suggests general agreement with predictions of the three-times rule. We define the coalescence ratio as the mitochondrial branch length for a species divided by intraspecific mtDNA diversity. We show that species with high coalescence ratios show nuclear monophyly, whereas species with low ratios have polyphyletic nuclear gene trees. As expected, species with intermediate coalescence ratios show a variety of patterns. Especially at very high or low coalescence ratios, the threetimes rule predicts nuclear gene patterns that can help detect the action of selection. The three-times rule may be useful as an empirical benchmark for evaluating evolutionary processes occurring at multiple loci. The application of molecular sequence data to systematics, population biology, and forensic identification of species often depends on whether monophyly has evolved at particular loci between taxa (Hudson 1992;Moritz 1994;Moore 1995;Palumbi and Cipriano 1998). Monophyly of genes is basic to some views of the species concept (Baum and Shaw 1995) and has been used to define evolutionarily significant units for management of threatened species (Moritz 1994). A population that has been separate long enough to develop gene monophyly has probably developed novel combinations of alleles at many genetic loci and may possess unique evolutionary features including adaptation to local environment that should be considered by resource managers (Moritz 1994). The shape of intra-and interspecific gene trees has also been used to infer the demographic history of species (Takahata 1995) and speciation patterns (M. P. Hare, F. Cipriano, and S. Palumbi, unpubl. ms.). Monophyly is also important if molecular tools are used in the identification of managed species in commercial markets (whales: Baker et al. 1993;Cipriano and Palumbi 1999; seals: Malik et al. 1997; canned fish: Rehbein et al. 1997; cannabis: Jagadish et al. 1996; caviar: Cohen 1997). This is...

Taxonomy is an imprecise science that delimits the evolutionary continuum into discrete categories. For marine mammals, this science is complicated by the relative lack of morphological data for taxa that inhabit remote and often vast ranges. We provide guidelines to promote consistency in studies relying primarily on molecular genetic data to delimit cetacean subspecies from both populations and species. These guidelines identify informational needs: basis for the taxonomic hypothesis being tested, description of current taxonomy, description of relevant life history, sample distribution, sample size, number and sequence length of genetic markers, description of measures taken to ensure data quality, summary statistics for the genetic markers, and analytical methods used to evaluate the genetic data. We propose an initial set of quantitative and qualitative standards based on the types of data and analytical methods most readily available at present. These standards are not expected to be rigidly applied. Rather, they are meant to encourage taxonomic arguments that are consistent and transparent. We hope professional societies, such as the Society for Marine Mammalogy, will adopt quantitative standards that evolve as new data types and analytical methods become widely available.

Coalescence theory predicts when genetic drift at nuclear loci will result in fixation of sequence differences to produce monophyletic gene trees. However, the theory is difficult to apply to particular taxa because it hinges on genetically effective population size, which is generally unknown. Neutral theory also predicts that evolution of monophyly will be four times slower in nuclear than in mitochondrial genes primarily because genetic drift is slower at nuclear loci. Variation in mitochondrial DNA (mtDNA) within and between species has been studied extensively, but can these mtDNA data be used to predict coalescence in nuclear loci? Comparison of neutral theories of coalescence of mitochondrial and nuclear loci suggests a simple rule of thumb. The "three-times rule" states that, on average, most nuclear loci will be monophyletic when the branch length leading to the mtDNA sequences of a species is three times longer than the average mtDNA sequence diversity observed within that species. A test using mitochondrial and nuclear intron data from seven species of whales and dolphins suggests general agreement with predictions of the three-times rule. We define the coalescence ratio as the mitochondrial branch length for a species divided by intraspecific mtDNA diversity. We show that species with high coalescence ratios show nuclear monophyly, whereas species with low ratios have polyphyletic nuclear gene trees. As expected, species with intermediate coalescence ratios show a variety of patterns. Especially at very high or low coalescence ratios, the three-times rule predicts nuclear gene patterns that can help detect the action of selection. The three-times rule may be useful as an empirical benchmark for evaluating evolutionary processes occurring at multiple loci.

We report the methods and results of molecular genetic identification of the species and, in some cases, geographical origins of whale and dolphin products purchased from retail markets and restaurants in Japan and South Korea. As reported previously (Baker & Palumbi 1994), we used the polymerase chain reaction (PCR) and a portable laboratory to amplify, purify and later sequence a portion of the mitochondrial DNA control region from 16 commercial products purchased in Japan. This ‘spot check’ revealed a surprising variety of species for sale, including minke, fin and humpback whales and one or two species of dolphins sold as ‘kujira’ or whale. In the Korean survey, DNA amplifications were conducted by two of us (C.S.B. and F.C.) working with independent equipment and reagents. The two sets of DNA amplifications were returned to our respective laboratories and sequenced independently for cross‐validation. Among the total of 17 species‐specific sequences we found a dolphin, a beaked whale, 13 Northern Hemisphere minke whales (representing at least seven distinct individuals) and two whales which are closely related to the recognized sei and Bryde's whales but could not be identified as either using available type sequences. We suggest that these two specimens represent a currently unrecognized species or subspecies of Bryde's whale, possibly the so‐called ‘small‐form’ reported from the tropical waters of the Indo‐Pacific. We conclude that molecular systematic analyses of DNA sequences have tremendous utility for the identification of whale and dolphin products. However, there are certain constraints on the application of these techniques for monitoring whaling or trade in whale products. First, PCR and DNA sequencing can generate misleading artefacts. These can generally be recognized or eliminated through experimental controls. Second, phylogenetic reconstructions of DNA sequences can be misinterpreted if the database of type sequences is inadequate or the taxonomy of the group is incomplete. This constraint is, at present, a more serious obstacle to molecular monitoring of whaling. Our results highlight uncertainties about the taxonomic status of oceanic populations and morphological forms of two species (or species complexes) targeted by legal and illegal hunting, the minke and Bryde's whales. Despite these uncertainties, it is difficult to reconcile some of the species available in Japanese and Korean commercial markets with recent catch records made available to the International Whaling Commission. It is particularly disturbing that two specimens of an unrecognized species or subspecies of baleen whale were for sale in a restaurant in South Korea in October, 1994, 8 years after the acceptance of an international moratorium on commercial whaling.