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PubMed2001

Genetic markers substantiate long-term storage and utilization of sperm by female painted turtles.

Most studies of genetic parentage in natural populations have been limited to a single breeding season or reproductive episode and, thus, provide only a snapshot of individuals' mating behaviours. Female turtles can store viable sperm in their reproductive tracts for as long as several years, but the extent to which this capacity is utilized in nature has remained unknown. Here, we employ microsatellite markers to assess genetic paternity in successive clutches of individually marked, free-ranging female painted turtles (Chrysemys picta) over a four year period. The genetic data from 113 clutches from this natural population demonstrate that most females (80.5%) remated each year and that each female generally used a single male's sperm to fertilize all clutches laid within a year. However, sperm usage among females varied considerably, and some females apparently used sperm that had been stored for up to three years to fertilize some or all eggs laid in consecutive nesting seasons. Thus, remating by females is not necessary for continued offspring production from a given sire. Furthermore, 13.2% of all clutches examined showed evidence of multiple paternity, and the genetic paternity patterns across years suggest a 'last in, first out' operation of the females' sperm storage tubules.

Alleles↗
PubMed1998

Single paternity of clutches and sperm storage in the promiscuous green turtle (Chelonia mydas).

Paternity of 22 green turtle (Chelonia mydas) clutches from 13 females of the southern Great Barrier Reef breeding population was determined through microsatellite analyses at five loci, including the analysis of successive clutches for nine of the females. A large number of alleles per locus (10-40) provided probabilities of detecting multiple paternity that were quite high, particularly at all loci combined (99.9%). Although green turtles are promiscuous breeders and there was an expectation of finding extensive multiple paternity, only two clutches were multiply sired and, in these, very few eggs had been fertilized by a secondary male. The rarity of multiple paternity may reflect either a low proportion of multiple matings by females in this population, or sperm competition, possibly resulting from a first-male sperm preference. Additionally, the analysis of > 900 offspring provided data on mutations, which included 20 mutation events that were observed in 27 offspring and involved both maternal and paternal lineages. Most mutations (n = 16) occurred at a single highly variable locus and their presence emphasizes the need to use multiple loci in paternity studies.

Animals↗
PubMed1999

A preliminary examination of genetic variation in a peripheral population of Blanding's turtle, Emydoidea blandingii.

Random amplified polymorphic DNA (RAPD) was used to compare the Nova Scotia population of Blanding's turtle (Emydoidea blandingii) with several populations from the species' main range. The Nova Scotia population is believed to have been isolated from the main range for 4000-8000 years. Cluster analysis using a neighbour-joining algorithm produced a dendrogram showing the Nova Scotia population clustering separately from those populations in the main range. Analysis of molecular variance shows 34.28% of total variance to be accounted for between the Nova Scotia population and populations in the main range. While this study is preliminary, the results suggest that the Nova Scotia population of Blanding's turtle may be important to the maintenance of genetic diversity in the species.

Analysis of Variance↗
PubMed2002

The leatherback turtle, Dermochelys coriacea, exhibits both polyandry and polygyny.

The leatherback turtle (Dermochelys coriacea) is an endangered species, and world-wide populations are declining. To understand better the mating structure of this pelagic and fragile species, we investigated paternity in nearly 1000 hatchlings from Playa Grande in Parque Marino Nacional Las Baulas, Costa Rica. We collected DNA samples from 36 adult female leatherbacks and assessed allele frequency distributions for three microsatellite loci. For 20 of these 36 females, we examined DNA from hatchlings representing multiple clutches, and in some cases assessed up to four successive clutches from the same female. We inferred paternal alleles by comparing maternal and hatchling genotypes. We could not reject the null hypothesis of single paternity in 12 of 20 families (31 of 50 clutches), but we did reject the null hypothesis in two families (eight of 50 clutches). In the remaining six families, the null hypothesis could not be accepted or rejected with certainty because the number of hatchlings exhibiting extra nonmaternal alleles was small, and could thus be a result of mutation or sample error. Successive clutches laid by the same female had the same paternal allelic contribution, indicating sperm storage or possibly monogamy. None of 20 females shared the same three-locus genotype whereas there were two instances of shared genotypes among 17 inferred paternal three-locus genotypes. We conclude that both polyandry and polygyny are part of the mating structure of this leatherback sea turtle population.

Animals↗
PubMed2000

Dedifferentiation of intrinsic response properties of motoneurons in organotypic cultures of the spinal cord of the adult turtle.

Explant cultures from the spinal cord of adult turtles were established and used to study the sensitivity of the intrinsic response properties of motoneurons to the changes in connectivity and milieu imposed by isolation in culture. Transverse sections 700 microm thick were explanted on cover slips and maintained in roller-tube cultures in medium containing serum and the growth factors brain-derived neurotrophin factor (BDNF), neurotrophin-3 (NT3), glial cell line-derived neurotrophic factor (GDNF) and ciliary neurotrophic factor (CNTF). The gross morphology of acute sections was maintained after 4 weeks in culture. Cell bodies of motoneurons remained stainable in fixed cultures with an antibody against choline acetyltransferase (ChAT) throughout the culture period. During culture, motoneurons maintained stable resting membrane potentials and were contacted by functional synapses. The ability to generate action potentials was also preserved as was delayed inward rectification and generation of calcium spikes in the presence of tetra-ethyl ammonium (TEA). In response to depolarization, however, motoneurons presented strong outward rectification, and only 41% of the cells recorded from maintained the ability to fire repetitively. By the second week in culture, a fraction of motoneurons displayed fast and slow transient outward rectification and low-threshold calcium spikes, features not seen in turtle motoneurons in acute slices. On the other hand, properties mediated by L-type Ca2+ channels disappeared during the first few days in culture. Our observations show that the phenotypical intrinsic response properties of mature spinal motoneurons are modified in explant cultures. The properties acquired resemble the properties in juvenile motoneurons in several species of terrestrial vertebrates.

Action Potentials↗
PubMed2000

Inositol 1,4,5-trisphosphate and adenophostin analogues induce responses in turtle olfactory sensory neurons.

Using the whole-cell mode of the patch-clamp technique, we recorded inward currents in response to inositol-1,4,5-trisphosphate (IP3) and adenophostin analogues in turtle olfactory sensory neurons. Dialysis of IP3 into the neurons induced inward currents with an increase in membrane conductance in a dose-dependent manner under the voltage-clamp conditions (holding potential -70 mV). The application of Ca2+-free Ringer solution to neurons previously dialysed with IP3 induced inward currents that were reversibly inhibited by application of Na+, Ca2+-free Ringer solution, normal Ringer solution or 10 microM ruthenium red. Dialysis of the adenophostin analogues, novel IP3 receptor ligands, also induced inward currents with an increase in membrane conductance. The magnitude of the responses to the adenophostin analogues varied among these derivatives. The application of Ca2+-free Ringer solution to neurons previously dialysed with the adenophostin analogues induced inward currents that were inhibited by the application of normal Ringer solution. The reversal potential of inward currents induced by an adenophostin analogue was similar to that induced by IP3, suggesting that inward currents induced by the adenophostin analogue were generated by a similar ionic mechanism to that induced by IP3. The present study demonstrated that IP3-mediated transduction pathways exist in turtle olfactory receptor neurons and that adenophostin analogues act as agonists of IP3.

Action Potentials↗
PubMed2001

The role of early neural activity in the maturation of turtle retinal function.

In the developing vertebrate retina, ganglion cells fire spontaneous bursts of action potentials long before the eye becomes exposed to sensory experience at birth. These early bursts are synchronised between neighbouring retinal ganglion cells (RGCs), yielding unique spatiotemporal patterns: 'waves' of activity sweep across large retinal areas every few minutes. Both at retinal and extraretinal levels, these embryonic retinal waves are believed to guide the wiring of the visual system using hebbian mechanisms of synaptic strengthening. In the first part of this review, we recapitulate the evidence for a role of these embryonic spontaneous bursts of activity in shaping developing complex receptive field properties of RGCs in the turtle embryonic retina. We also discuss the role of visual experience in establishing RGC visual functions, and how spontaneous activity and visual experience interact to bring developing receptive fields to maturation. We have hypothesised that the physiological changes associated with development reflect modifications in the dendritic arbours of RGCs, the anatomical substrate of their receptive fields. We demonstrate that there is a temporal correlation between the period of receptive field expansion and that of dendritic growth. Moreover, the immature spontaneous activity contributes to dendritic growth in developing RGCs. Intracellular staining of RGCs reveals, however, that immature receptive fields only rarely show direct correlation with the layout of the corresponding dendritic tree. To investigate the possibility that not only the presence of the spontaneous activity, but even the precise spatiotemporal patterns encoded in retinal waves might contribute to the refinement of retinal neural circuitry, first we must clarify the mechanisms mediating the generation and propagation of these waves across development. In the second part of this review, we present evidence that turtle retinal waves, visualised using calcium imaging, exhibit profound changes in their spatiotemporal patterns during development. From fast waves sweeping across large retinal areas and recruiting many cells on their trajectory at early stages, waves become slower and eventually stop propagating towards hatching, when they become stationary patches of neighbouring coactive RGCs. A developmental switch from excitatory to inhibitory GABAA responses appears to mediate the modification in spontaneous activity patterns while the retina develops. Future chronic studies using specific spatiotemporal alterations of the waves will shed a new light on how the wave dynamics help in sculpting retinal receptive fields.

Action Potentials↗
PubMedDate not supplied

Morphogenesis of the turtle shell: the development of a novel structure in tetrapod evolution.

The turtle shell is an evolutionary novelty that is synapomorphic for chelonians. The carapace is initiated by the entrapment of the ribs by the carapacial ridge (CR), a lateral bulge of the dorsal ectoderm and dermal mesoderm. The mechanisms by which the CR is initiated, the ribs entrapped and the dorsal dermis ossified, remains unknown. Similarly, the formation of the plastron remains unexplained. Here, we present a series of anatomical investigations into plastron and carapace formation in the red-eared slider, Trachemys scripta, and the snapping turtle, Chelydra serpentina. We document the entrapment of the ribs by the CR and the formation of the plastron and carapacial bones by intramembranous ossification. We note the formation of the ossification centers around each rib, which suggest that the rib is organizing dermal ossification by secreting paracrine factors. The nuchal ossification center is complex and appears to involve multiple bone-forming regions. Individual ossification centers at the periphery of the carapace form the peripheral and pygial bones. The intramembranous ossification of the plastron proceeds from nine distinct ossification centers, and there appear to be interactions between the spicules of apposing centers as they draw near each other.

Animal Structures↗
PubMed2005

Two new species of Eimeria (Apicomplexa: Eimeriidae) from Asian geoemydid turtles Kachuga tentoria and Melanochelys trijuga (Testudines: Geoemydidae).

Coprological examination of ten Pink-ringed tent turtles Kachuga tentoria circumdata, recently imported from India, and three Burmese black turtles Melanochelys trijuga edeniana, imported from Myanmar, revealed the presence of two new species of Eimeria. Oocysts of Eimeria kachua n. sp. from K. t. circumdata are broadly oval to subspherical, 15.3 (13-18) x 13.9 (12-16) microm, with polar granule and subspherical oocyst residuum. Sporocyst elongatelly oval to spindle-shaped, 8.7 (7.5-10) x 4.9 (4-6) microm, with a knoblike Stieda body, covered with fine membranous cupola-like structures. Thin walled oocysts of Eimeria patta n. sp. from M. t. edeniana, have an irregular shape, influenced by the position of sporocysts, frequently with lobular irregularities, 12.6 (11-16) x 9.1 (7.5-12) microm. Sporocysts are oval to ellipsoidal, 5.8 (5-7) x 4.2 (3.5-5) microm.

Animals↗
PubMedDate not supplied

Interactions of Salmonella enterica subsp. enterica serovar Muenchen with intestinal explants of the turtle Trachemys scripta scripta.

Salmonella infections in reptiles, in contrast to those in birds and mammals, are limited to the intestinal tract. In this study, interactions of a strain of Salmonella enterica subsp. enterica serovar Muenchen (SEEM) with intestinal explants of the turtle Trachemys scripta scripta were examined by scanning electron microscopy (SEM). Adhesion and invasion in the chelonian intestinal explants at 30 degrees C and 37 degrees C were evaluated quantitatively. For purposes of comparison, the invasive capacity of SEEM in the continuous avian epithelial cell line DIV-1 at 30 degrees C and 37 degrees C was determined. Small numbers of M-like cells were found in the ileum of the turtles. The bacteria adhered mainly to the mucus of the intestinal explants. Only small numbers of salmonellae were associated with epithelial cells. Higher numbers of bacteria adhered at 30 degrees C than at 37 degrees C. Epithelial damage, embedding of bacteria in the epithelial surface and a ruffling-like process were noted only at 37 degrees C. Minimal numbers of salmonellae invaded the explants at 30 degrees C and 37 degrees C. Invasion of DIV-1 cells was greater at 37 degrees C than at 30 degrees C. The study suggested that the intestinal mucous layer provides an important site of colonization for salmonellae in the chelonian host and protects the underlying epithelial cells.

Animal Diseases↗
PubMed1972

Biosynthesis of acetylcholine in turtle photoreceptors.

For determination of possible neurotransmitters synthesized by photoreceptor cells, turtle retinas were dissociated into single cells with proteolytic enzymes. These cells were partially separated by velocity sedimentation to yield a fraction rich in photoreceptors. Individual photoreceptor cells were then sucked into a micropipette and incubated with labeled precursors of known or suspected neurotransmitters. After incubation, the radioactive products were analyzed by high-voltage electrophoresis. Of all the chemicals tested, turtle photoreceptor cells synthesized only acetylcholine, suggesting that these cells may be cholinergic.

Acetylcholine↗
PubMed1979

Effects of pseudomonas toxin A, diphtheria toxin, and cholera toxin on electrical characteristics of turtle bladder.

Rapidly developing changes in the short-circuiting current (Isc), conductance (G), and potential (PD) of turtle bladders in Na-rich or Na-free media are seen after the mucosal addition, at 10 nM, of each of three toxins that contain ADP-ribosylation activity: Pseudomonas aeruginosa toxin A, diphtheria toxin, and cholera toxin. Toxin A irreversibility decreased the Isc, PD, and G of bladders in Na-rich media and the Isc and PD of bladders in Na-free media. Diphtheria or cholera toxin reversibly increased Isc and PD (not G), but only in Na-free media. The effects of toxin A in the turtle bladder, like those in other host cell systems, were eliminated by preexposure of this toxin to heat, specific antitoxin, or dithiothreitol and urea. Because exposure to this last condition increases the ADP-ribosylation activity of toxin A, it is suggested that the proenzyme is the required transport-inhibiting form of toxin A. The effects of all three toxins occurred rapidly, possibly before any of the possible intracellular ADP-ribosylation reactions are initiated. Whereas a recognition binding of toxin of toxin to receptors on the apical membrane completely accounts for the reversible effects of diphtheria or cholera toxin, this and additional toxin-membrane interactions (e.g., translocation) are needed to account for the irreversible effects of toxin A.

Animals↗
PubMed1981

Involvement of small-field horizontal cells in feedback effects on green cones of turtle retina.

Light stimuli depolarize green cones of turtle retina through a circuit involving a feedback connection from luminosity horizontal cells (L-HC) to green cones. In turtle retina two types of L-HC have been distinguished: large-field L-HC and small-field L-HC. The spatial properties of the feedback depolarizations of green cones were compared with those of both large- and small-field L-HC. Green cones were found to be more effectively depolarized by relatively small spots of red light than by large red annuli. Moreover, red light stimulation of the periphery of the receptive field could reduce the depolarizing influence of central red stimuli. These spatial properties greatly differ from those of the large-field L-HC, whereas they strongly resemble those of the small-field L-HC. These results suggest that the small-field L-HC mediate the feedback action on green cones.

Animals↗
PubMed1982

Exocytosis regulates urinary acidification in turtle bladder by rapid insertion of H+ pumps into the luminal membrane.

Urinary acidification by the turtle bladder is due to a H+-ATPase that is located in the luminal membrane. The rate of H+ transport is stimulated by an increase in the ambient CO2. Using the fluorescent dye acridine orange, we showed that the mitochondria-rich cell of this equilibrium contains vesicles whose internal pH is acidic. We measured the pH of these vesicles by using endocytosed fluorescein isothiocyanate-labeled dextran and found it to be near 5.0. The pH increased after treatment with protonophores or metabolic inhibitors, suggesting that it was due to a H+ pump rather than to a Donnan effect. In bladders preloaded with fluorescent dextran, CO2 stimulated exocytosis and H+ transport measured simultaneously in the same bladder. The increase in the H+ current correlated well with the extent of exocytosis, and both were inhibited by pretreatment with colchicine. We conclude that the turtle bladder contains an intracellular reserve of vesicles containing H+ pumps and CO2 stimulates rapid fusion of these vesicles with the luminal membrane with consequent insertion of H+ pumps, thereby stimulating H+ secretion across the whole epithelium.

Acid-Base Equilibrium↗
PubMed1993

A molecular phylogeny for marine turtles: trait mapping, rate assessment, and conservation relevance.

Nucleotide sequences from the cytochrome b gene of mitochondrial DNA were employed to resolve phylogenetic controversies and to assess molecular evolutionary rates in marine turtles (Chelonioidea). Findings of special relevance to conservation biology include discovery of a distant relationship between Natator and other cheloniid species, the paraphyly of Chelonia mydas with respect to Chelonia agassizi, and genetic distinctiveness of Lepidochelys kempi from Lepidochelys olivacea. A longstanding debate in evolutionary ecology was resolved by phylogenetic mapping of dietary habits, which indicates that the spongivore Eretmochelys imbricata evolved from a carnivorous rather than a herbivorous ancestor. Sequence divergences at intergeneric and interfamilial levels, when assessed against fossil-based separation times, support previous suggestions (from microevolutionary comparisons) that mitochondrial DNA in marine turtles evolves much more slowly than under the "conventional" vertebrate clock. This slow pace of nucleotide replacement is consistent with recent hypotheses linking substitution rate to generation length and metabolic pace.

Animals↗
PubMed1989

Excretion and metabolism of a series of xanthines by the turtle Pseudemys scripta elegans.

The xanthines paraxanthine, xanthine, and uric acid are absorbed completely by the turtle Pseudemys scripta elegans and metabolised presumably into CO2 and NH3. No intermediate metabolites are excreted. Caffeine, hydroxycaffein, theobromine, hydroxytheobromine, and hydroxypraxanthine are absorbed and excreted in different amounts or percentages of the dose administered. Intermediate metabolites of these compounds are not excreted. The turtle Pseudemys scripta elegans may, therefore, be ammonotelic.

Animals↗