Vocal exchanges of signature whistles in bottlenose dolphins (<Emphasis Type="Italic">Tursiops truncatus</Emphasis>)

Bottlenose dolphins (Tursiops truncatus) produce individually distinctive vocalizations—referred to as “signature whistles”—that are thought to function as an individual and conspecific recognition system for maintenance of consistent contact between individuals. Observations and playback experiments were conducted at aquariums to study these whistle–vocal exchanges in bottlenose dolphins. Temporal patterns of vocalization were examined by analyzing the intercall intervals between two consecutive whistles. When a second individual produced a call that was different from the first individual’s vocalization, most of these calls were shorter than 1 s. However, when two consecutive calls were produced by the same individual, the second call rarely occurred within 1 s of the first. These results suggest that a second whistle may be produced by a different caller in response to the first whistle; however, in the case of an absence of a response, the first caller is likely to give further whistles. The results of this acoustic analysis show that the dolphins used in this study mostly used signature whistles during the recorded vocal exchanges.     

The development of diving bradycardia in bottlenose dolphins (<Emphasis Type="Italic">Tursiops truncatus</Emphasis>)

Bradycardia is an important component of the dive response, yet little is known about this response in immature marine mammals. To determine if diving bradycardia improves with age, cardiac patterns from trained immature and mature bottlenose dolphins (Tursiops truncatus) were recorded during three conditions (stationary respiration, voluntary breath-hold, and shallow diving). Maximum (mean: 117±1 beats·min^–1) and resting (mean: 101±5 beats·min^–1) heart rate (HR) at the water surface were similar regardless of age. All dolphins lowered HR in response to apnea; mean steady state breath-hold HR was not correlated with age. However, the ability to reduce HR while diving improved with age. Minimum and mean steady state HR during diving were highest for calves. For example, 1.5–3.5-year-old calves had significantly higher mean steady state diving HR (51±1 beats·min^–1) than 3.5–5.5-year-old juveniles (44±1 beats·min^–1). As a result, older dolphins demonstrated greater overall reductions in HR during diving. Longitudinal studies concur; the ability to reduce HR improved as individual calves matured. Thus, although newly weaned calves as young as 1.7 years exhibit elements of cardiac control, the capacity to reduce HR while diving improves with maturation up to 3.5 years postpartum. Limited ability for bradycardia may partially explain the short dive durations observed for immature marine mammals.Abbreviations ADL aerobic dive limit - cADL calculated aerobic dive limit - ECG electrocardiogram - HR heart rate - TDR time–depth recorder Communicated by L.C.-H. Wang     

Seasonal patterns of heat loss in wild bottlenose dolphins (<Emphasis Type="Italic">Tursiops truncatus</Emphasis>)

This study investigated patterns of heat loss in bottlenose dolphins (Tursiops truncatus) resident to Sarasota Bay, FL, USA, where water temperatures vary seasonally from 11 to 33°C. Simultaneous measurements of heat flux (HF) and skin surface temperature were collected at the body wall and appendages of dolphins during health-monitoring events in summer (June 2002–2004) and winter (February 2003–2005). Integument thickness was measured and whole body conductance (W/m² °C) was estimated using HF and colonic temperature measurements. Across seasons, HF values were similar at the appendages, but their distribution differed significantly at the flipper and fluke. In summer, these appendages displayed uniformly high values, while in winter they most frequently displayed very low HF values with a few high HF values. In winter, blubber thickness was significantly greater and estimated conductance significantly lower, than in summer. These results suggest that dolphins attempt to conserve heat in winter. In winter, though, HF values across the body wall were similar to (flank) or greater than (caudal keel) summer values. It is likely that higher winter HF values are due to the steep temperature gradient between the body core and colder winter water, which may limit the dolphin’s ability to decrease heat loss across the body wall.     

Modulation rate transfer functions in bottlenose dolphins (<Emphasis Type="Italic">Tursiops truncatus</Emphasis>) with normal hearing and high-frequency hearing loss

Envelope following responses were measured in two bottlenose dolphins in response to sinusoidal amplitude modulated tones with carrier frequencies from 20 to 60 kHz and modulation rates from 100 to 5,000 Hz. One subject had elevated hearing thresholds at higher frequencies, with threshold differences between subjects varying from ±4 dB at 20 and 30 kHz to +40 dB at 50 and 60 kHz. At each carrier frequency, evoked response amplitudes and phase angles were plotted with respect to modulation frequency to construct modulation rate transfer functions. Results showed that both subjects could follow the stimulus envelope components up to at least 2,000 Hz, regardless of carrier frequency. There were no substantial differences in modulation rate transfer functions for the two subjects suggesting that reductions in hearing sensitivity did not result in reduced temporal processing ability. In contrast to earlier studies, phase data showed group delays of approximately 3.5 ms across the tested frequency range, implying generation site(s) within the brainstem rather than the periphery at modulation rates from 100 to 1,600 Hz. This discrepancy is believed to be the result of undersampling of the modulation rate during previous phase measurements.     

Effect of valuable relationship on reconciliation and initiator of reconciliation in captive bottlenose dolphins (<Emphasis Type="Italic">Tursiops truncatus</Emphasis>)

Reconciliation, post-conflict affiliation between former opponents, was suggested to repair the relationship between former opponents. The number of times that each former opponent initiates reconciliation may be affected by the dominance style. Despotic dominance style was suggested to restrain the initiation of reconciliation by recipients of aggression, unlike relaxed dominance style. Although reconciliation was suggested to restore relationships, reconciliation sometimes did not occur. The valuable relationship hypothesis predicts that reconciliation is more likely to occur when former opponents share a valuable relationship. However, few studies have tested this hypothesis in non-primate species. This study investigated, in captive bottlenose dolphins, which are suggested as having a relaxed dominance style, whether aggressors or recipients of aggression initiate reconciliation, and whether relationships between former opponents affect the occurrence of reconciliation. The number of times that aggressors or recipients of aggression initiate reconciliation did not differ. Recipients of aggression initiated reconciliation sooner than aggressors. These results support the prospect that recipients of aggression are not impeded in initiating reconciliation in relaxed species. Former opponents who share a more affiliative relationship were reconciled more frequently. Bottlenose dolphins were suggested to receive the benefits from individuals who shared more affiliative relationships. The valuable relationship hypothesis may be supported in bottlenose dolphins.     

Mitochondrial and nuclear DNA analyses reveal fine scale geographic structure in bottlenose dolphins (<Emphasis Type="Italic">Tursiops truncatus</Emphasis>) in the Gulf of Mexico

There is a need for biological information to support current stock designations of bottlenose dolphins (Tursiops truncatus) in the Gulf of Mexico. The existence of many inshore, resident “communities” raises questions as to the relationship these dolphins may hold with dolphins inhabiting neighboring inshore and coastal areas. In this study, population subdivision was examined among four resident, inshore bottlenose dolphin stocks (Sarasota Bay, FL, Tampa Bay, FL, Charlotte Harbor, FL and Matagorda Bay, TX) and one coastal stock (1–12 km offshore) in the Gulf of Mexico. Evidence of significant population structure among all areas was found on the basis of both mitochondrial DNA (mtDNA) control region sequence data and nine nuclear microsatellite loci. Estimates of relatedness showed no population contained a significantly high number of related individuals, while separate AMOVAs for males and females indicated that both sexes exhibit a significant level of site philopatry. Results presented here provide the first genetic evidence of population subdivision between the coastal Gulf of Mexico and adjacent inshore areas along the central west coast of Florida. Such strong genetic subdivision is surprising given the short geographical distance between many of these areas and the lack of obvious geographic barriers to prevent gene flow. These findings support the current, separate identification of stocks for bottlenose dolphins inhabiting the eastern coastal and inshore areas of the Gulf of Mexico.     

New combinations and typifications in <Emphasis Type="Italic">Bistorta</Emphasis>, <Emphasis Type="Italic">Persicaria</Emphasis>, <Emphasis Type="Italic">Polygonum,</Emphasis> and <Emphasis Type="Italic">Rumex</Emphasis> (Polygonaceae)

New combinations are proposed in anticipation of the Polygonaceae treatment in the forthcoming volume of Intermountain Flora: Polygonum kelloggii var. esotericum, P. kelloggii var. watsonii , Rumex densiflorus var. pycnanthus , R. salicifolius var. utahensis, and R. occidentalis var. tomentellus. Typifications are proposed to facilitate ongoing studies in Polygonaceae and to maintain current usage.     

Environmental and social influences on the genetic structure of bottlenose dolphins (<Emphasis Type="Italic">Tursiops aduncus</Emphasis>) in Southeastern Australia

Determining genetic connectivity of bottlenose dolphin communities helps identify evolutionary mechanisms, such as environmental and social factors, that interact to shape dispersal in highly social marine mammals. Here, we expand on a localized study that found marked genetic differentiation among resident dolphins (Tursiops aduncus) in the Port Stephens embayment and adjacent coastal communities, to include four additional communities inhabiting different environment types along the New South Wales coast, Southeastern Australia. Analysis of the mitochondrial DNA control region and seven microsatellite loci suggest the nine communities may have originated from a single ancestral population that progressively colonised the coast in a southward direction. Gene flow among communities was predominately governed by habitat type. The two enclosed embayments showed the highest level of genetic differentiation from other communities, while genetic differentiation among coastal and open embayment communities generally followed a pattern of isolation by distance. Directional bias in recent migration rates was evident, with the centrally located Hunter coast communities consisting of individuals with mixed ancestry from the Northern, Southern and Port Stephens communities. Emigration from Port Stephens was substantially higher than in the opposite direction, indicating there may be social barriers to dispersal created by Port Stephens dolphins. Our results suggest that the scale of connectivity of bottlenose dolphin communities inhabiting heterogeneous environments is likely to be affected by local habitat adaptation. This has important implications for the management of communities exposed to increasing levels of anthropogenic disturbances, such as the intensive commercial dolphin-watching industry operating in Port Stephens.     

<Emphasis Type="Italic">Ty</Emphasis>1<Emphasis Type="Italic">/copia</Emphasis>- and <Emphasis Type="Italic">Ty</Emphasis>3<Emphasis Type="Italic">/gypsy</Emphasis>-like DNA sequences in <Emphasis Type="Italic">Helianthus </Emphasis>species

Two repeated DNA sequences isolated from a partial genomic DNA library of Helianthus annuus, p HaS13 and p HaS211, were shown to represent portions of the int gene of a Ty3 /gypsy retroelement and of the RNase-Hgene of a Ty1 /copia retroelement, respectively. Southern blotting patterns obtained by hybridizing the two probes to BglII- or DraI-digested genomic DNA from different Helianthus species showed p HaS13 and p HaS211 were parts of dispersed repeats at least 8 and 7 kb in length, respectively, that were conserved in all species studied. Comparable hybridization patterns were obtained in all species with p HaS13. By contrast, the patterns obtained by hybridizing p HaS211 clearly differentiated annual species from perennials. The frequencies of p HaS13- and p HaS211-related sequences in different species were 4.3x10(4)-1.3x10(5) copies and 9.9x10(2)-8.1x10(3) copies per picogram of DNA, respectively. The frequency of p HaS13-related sequences varied widely within annual species, while no significant difference was observed among perennial species. Conversely, the frequency variation of p HaS211-related sequences was as large within annual species as within perennials. Sequences of both families were found to be dispersed along the length of all chromosomes in all species studied. However, Ty3 /gypsy-like sequences were localized preferentially at the centromeric regions, whereas Ty1/ copia-like sequences were less represented or absent around the centromeres and plentiful at the chromosome ends. These findings suggest that the two sequence families played a role in Helianthusgenome evolution and species divergence, evolved independently in the same genomic backgrounds and in annual or perennial species, and acquired different possible functions in the host genomes.     

<Emphasis Type="Italic">Quinua</Emphasis> and Relatives (<Emphasis Type="Italic">Chenopodium</Emphasis> sect.<Emphasis Type="Italic">Chenopodium</Emphasis> subsect.<Emphasis Type="Italic">Celluloid</Emphasis>)

Traditionally viewed as an Andean grain crop,Chenopodium quinoa Willd. includes domesticated populations that are not Andean, and Andean populations that are not domesticated. Comparative analysis of leaf morphology and allozyme frequencies have demonstrated that Andean populations, both domesticated(quinua) and free-living(ajara), represent an exceptionally homogeneous unit that is well differentiated from allied domesticates of coastal Chile(quingua) and freeliving populations of the Argentine lowlands(C. hircinum). This pattern of relationships indicates that Andean populations represent a monophyletic crop/weed system that has possibly developed through cyclic differentiation (natural vs. human selection) and introgressive hybridization. Relative levels of variation suggest that this complex originated in the southern Andes, possibly from wild types allied withC. hircinum, with subsequent dispersal north to Colombia and south to the Chilean coast. Coastal populations were apparently isolated from post-dispersal differentiation and homogenization that occurred in the Andes. Other data point toward a center of origin in the northern Andes with secondary centers of genetic diversity subsequently developing in the southern Andes and the plains of Argentina. Comparative linkage of South American taxa, all tetraploid, with North American tetraploids of the subsection will eventually clarify this problem. While the possibility of a direct phyletic connection betweenC. quinoa and the Mexican domesticate(C. berlandieri subsp. nuttalliae,) cannot be excluded, available evidence indicates that the latter represents an autonomous lineage that is associated with the basal tetraploid, C. b. subsp.berlandieri, through var.sinuatum, whereas South American taxa show possible affinities to either var. zschackei or var.berlandieri. An extinct domesticate of eastern North America,C. b. subsp.jonesianum, represents either another instance of independent domestication, possibly from subsp. b. var.zschackei, or a northeastern outlier of subsp.nuttalliae.     

Floral development and systematic position of <Emphasis Type="Italic">Arillastrum</Emphasis>, <Emphasis Type="Italic">Allosyncarpia</Emphasis>, <Emphasis Type="Italic">Stockwellia</Emphasis> and <Emphasis Type="Italic">Eucalyptopsis</Emphasis> (Myrtaceae)

We have investigated the floral ontogeny of Arillastrum, Allosyncarpia, Stockwellia and Eucalyptopsis (of the eucalypt group, Myrtaceae) using scanning electron microscopy and light microscopy. Several critical characters for establishing relationships between these genera and to the eucalypts have been determined. The absence of compound petaline primordia in Arillastrum, Allosyncarpia, Stockwellia and Eucalyptopsis excludes these taxa from the eucalypt clade. Post-anthesis circumscissile abscission of the hypanthium above the ovary in Stockwellia, Eucalyptopsis and Allosyncarpia is evidence that these three taxa form a monophyletic group; undifferentiated perianth parts and elongated fusiform buds are characters that unite Stockwellia and Eucalyptopsis as sister taxa. No floral characters clearly associate Arillastrum with either the eucalypt clade or the clade of Stockwellia, Eucalyptopsis and Allosyncarpia.We gratefully acknowledge Clyde Dunlop and Bob Harwood (Northern Territory Herbarium) for collecting specimens of Allosyncarpia, and Bruce Gray (Atherton) for collecting specimens of Stockwellia. The Australian National Herbarium (CANB) kindly lent herbarium specimens of Eucalyptopsis for examination. This research was supported by a University of Melbourne Research Development Grant to Andrew Drinnan.     

<Emphasis Type="Italic">Galleria</Emphasis><Emphasis Type="Italic">mellonella</Emphasis> are Resistant to <Emphasis Type="Italic">Pneumocystis</Emphasis><Emphasis Type="Italic">murina</Emphasis> Infection

Studying Pneumocystis has proven to be a challenge from the perspective of propagating a significant amount of the pathogen in a facile manner. The study of several fungal pathogens has been aided by the use of invertebrate model hosts. Our efforts to infect the invertebrate larvae Galleria mellonella with Pneumocystis proved futile since P. murina neither caused disease nor was able to proliferate within G. mellonella. It did, however, show that the pathogen could be rapidly cleared from the host.     

Revision of the <Emphasis Type="Italic">Serrulati</Emphasis> species of <Emphasis Type="Italic">Penstemon</Emphasis> section <Emphasis Type="Italic">Saccanthera</Emphasis> subsection <Emphasis Type="Italic">Serrulati</Emphasis>

A revision of Penstemon sect. Saccanthera subsect. Serrulati includes a new species (P. salmonensis), a new variety (P. triphyllus var. infernalis), and the elevation of a subspecies to species (P. curtiflorus), bringing the total number of species to eight, which are keyed and described, complete with nomenclature and type citations.     

Pedogamy in <Emphasis Type="Italic">Neidium</Emphasis> (<Emphasis Type="Italic">Bacillariophyceae</Emphasis>)

Single (unpaired) vegetative cells of freshwater pennate diatom Neidium cf. ampliatum differentiated into gametangia and produced a single zygote (auxospore) via a pedogamic process. The gametic nuclei fused after auxospore expansion had begun. The auxospore expanded in parallel to the apical axis of the gametangium.     

Sociality in <Emphasis Type="Italic">Euglossa</Emphasis> (<Emphasis Type="Italic">Euglossa</Emphasis>) <Emphasis Type="Italic">viridissima</Emphasis> Friese (Hymenoptera,Apidae, Euglossini)

Euglossa viridissima is an orchid bee that forms both solitary and multiple female nests, making it a suitable species for the study of factors leading to diverse degrees of sociality in Euglossines. We conducted observations in eight reused nests (where a first generation of bees had been produced) kept in artificial boxes from the Yucatan Peninsula, Mexico. Five nests were reused (reactivated) by a single female (SFN), two nests reused by a mother and one daughter (MFN1) and one nest reused by the mother and two daughters (MFN2). No single nest was reactivated by unrelated females. The number of foraging trips, their duration and the duration of cell provisioning was not different between SFN and MFN. The overall production of cells per female was not different either between both types of nest. However, in MFN although all females did lay eggs, there was a reproductive skew in favor of the mother (95 and 45% of the brood produced in MFN1 and MFN2 respectively). She showed reproductive control of her daughters through oophagy and displaying threatening behavior when the daughters tried to open a cell where she had laid an egg. Brood losses to parasites (Anthrax sp. (Bom-byliidae) and Hoplostelis bivittata (Megachilidae)) were only found in SFN which possibly reflects and advantage of MFN in this respect. Our results coupled with other studies in Euglossa, reveal that a wide range of social behaviors occur in this genus, from solitary and communal to primitive reproductive division of labor. Multiple factors involving different levels of pressure imposed by food availability and parasites may favor such a diverse range of nesting behaviors. Interestingly, female associations in E. viridissima seem a result of kin selection that is enforced by coercion from mother females on their daughters. More studies are needed to shed light upon the social organization of Euglossa and other Euglossines and on their phylogenetic relationships in order to trace the origins of eusociality in Apidae. Received 12 February 2008; revised 25 June 2008; accepted 17 July 2008.     

<Emphasis Type="Italic">Agrobacterium</Emphasis><Emphasis Type="Italic">tumefaciens</Emphasis>-mediated transformation of callus cells of <Emphasis Type="Italic">Crataegus</Emphasis><Emphasis Type="Italic">aronia</Emphasis>

A genetic transformation system has been developed for callus cells of Crataegus aronia using Agrobacterium tumefaciens. Callus culture was established from internodal stem segments incubated on Murashige and Skoog (MS) medium supplemented with 5 mg l⁻¹ Indole-3-butyric acid (IBA) and 0.5 mg l⁻¹ 6-benzyladenine (BA). In order to optimize the callus culture system with respect to callus growth and coloration, different types and concentrations of plant growth regulators were tested. Results indicated that the best average fresh weight of red colored callus was obtained on MS medium supplemented with 2 mg l⁻¹ 2,4-dichlorophenoxyacetic acid (2,4-D) and 1.5 mg l⁻¹ kinetin (Kin) (callus maintenance medium). Callus cells were co-cultivated with Agrobacterium harboring the binary plasmid pCAMBIA1302 carrying the mgfp5 and hygromycin phosphotransferase (hptII) genes conferring green fluorescent protein (GFP) activity and hygromycin resistance, respectively. Putative transgenic calli were obtained 4 weeks after incubation of the co-cultivated explants onto maintenance medium supplemented with 50 mg l⁻¹ hygromycin. Molecular analysis confirmed the integration of the transgenes in transformed callus. To our knowledge, this is the first time to report an Agrobacterium-mediated transformation system in Crataegus aronia.     

<Emphasis Type="Italic">In silico</Emphasis> and <Emphasis Type="Italic">in situ</Emphasis> characterization of the zebrafish (<Emphasis Type="Italic">Danio rerio</Emphasis>) <Emphasis Type="Italic">gnrh3</Emphasis> (sGnRH) gene

Background

Gonadotropin releasing hormone (GnRH) is responsible for stimulation of gonadotropic hormone (GtH) in the hypothalamus-pituitary-gonadal axis (HPG). The regulatory mechanisms responsible for brain specificity make the promoter attractive for in silico analysis and reporter gene studies in zebrafish (Danio rerio).

Results

We have characterized a zebrafish [Trp⁷, Leu⁸] or salmon (s) GnRH variant, gnrh 3. The gene includes a 1.6 Kb upstream regulatory region and displays the conserved structure of 4 exons and 3 introns, as seen in other species. An in silico defined enhancer at -976 in the zebrafish promoter, containing adjacent binding sites for Oct-1, CREB and Sp1, was predicted in 2 mammalian and 5 teleost GnRH promoters. Reporter gene studies confirmed the importance of this enhancer for cell specific expression in zebrafish. Interestingly the promoter of human GnRH-I, known as mammalian GnRH (mGnRH), was shown capable of driving cell specific reporter gene expression in transgenic zebrafish.

Conclusions

The characterized zebrafish Gnrh3 decapeptide exhibits complete homology to the Atlantic salmon (Salmo salar) GnRH-III variant. In silico analysis of mammalian and teleost GnRH promoters revealed a conserved enhancer possessing binding sites for Oct-1, CREB and Sp1. Transgenic and transient reporter gene expression in zebrafish larvae, confirmed the importance of the in silico defined zebrafish enhancer at -976. The capability of the human GnRH-I promoter of directing cell specific reporter gene expression in zebrafish supports orthology between GnRH-I and GnRH-III.

     

The tyrosine <Emphasis Type="Italic">O</Emphasis>-prenyltransferase SirD catalyzes <Emphasis Type="Italic">O</Emphasis>-, <Emphasis Type="Italic">N</Emphasis>-, and <Emphasis Type="Italic">C</Emphasis>-prenylations

Recently, the prenyltransferase SirD was found to be responsible for the O-prenylation of tyrosine in the biosynthesis of sirodesmin PL in Leptosphaeria maculans. In this study, the behavior of SirD towards phenylalanine/tyrosine and tryptophan derivatives was investigated. Product formation has been observed with 12 of 19 phenylalanine/tyrosine derivatives. It was shown that the alanine structure attached to the benzene ring and an electron donor, e.g., OH or NH₂, at its para-position are essential for the enzyme activity. Modifications were possible both at the side chain and the benzene ring. Enzyme products from seven phenylalanine/tyrosine derivatives were isolated and characterized by MS and NMR analyses including HSQC and HMBC and proven to be O- or N-prenylated derivatives at position C4 of the benzene rings. K _M values of six selected derivatives were found in the range of 0.10–0.68 mM. Catalytic efficiencies (K _cat/K _M ) were determined in the range of 430–1,110 s⁻¹·M⁻¹ with l-tyrosine as the best substrate. In addition, 7 of 14 tested tryptophan analogs were also accepted by SirD and converted to C7-prenylated derivatives, which was confirmed by comparison with products obtained from enzyme assays using a 7-dimethylallyltryptophan synthase 7-DMATS from Aspergillus fumigatus.     

Isolation and characterization of the <Emphasis Type="Italic">Larix gmelinii </Emphasis><Emphasis Type="Italic">ANGUSTIFOLIA</Emphasis> (<Emphasis Type="Italic">LgAN</Emphasis>) gene

ANGUSTIFOLIA (AN), a plant homolog of C-terminal binding protein, controls the polar elongation of leaf cells and the trichome-branching pattern in Arabidopsis thaliana. In the present study, degenerate PCR was used to isolate an ortholog of AN, referred to as LgAN, from larch (Larix gmelinii). The LgAN cDNA is predicted to encode a protein of 646 amino acids that shows striking sequence similarity to AN proteins from other plants. The predicted amino acid sequence has a conserved NAD-dependent 2-hydroxy acid dehydrogenase (D2-HDH) motif and a plant AN-specific LxCxE/D motif at its N-terminus, as well as a plant-specific long C-terminal region. The LgAN gene is a single-copy gene that is expressed in all larch tissues. Expression of the LgAN cDNA rescued the leaf width and trichome-branching pattern defects in the angustifolia-1 (an-1) mutant of Arabidopsis, showing that the LgAN gene has effects complementary to those of AN. These results suggest that the LgAN gene has the same function as the AN gene.