Microsatellite DNA loci from the Diamondback terrapin (Malaclemys terrapin)

We describe polymerase chain recation (PCR) primers and conditions to amplify one dinucleotide and five tetranucleotide microsatellite DNA loci isolated from the Diamondback terrapin (Malaclemys terrapin). The PCR primers were tested on 21 terrapins from Cape Romain, SC, USA. The microsatellite primers developed yielded a high number of alleles (8–14) and high observed heterozygosities (0.57–1.0).     

Evidence of Eelgrass (Zostera marina) Seed Dispersal by Northern Diamondback Terrapin (Malaclemys terrapin terrapin) in Lower Chesapeake Bay

The initial discovery in May 2009 of eelgrass (Zostera marina) seeds in fecal samples of wild-caught northern diamondback terrapins (Malaclemys terrapin terrapin) was the first field evidence of eelgrass seed ingestion in this species. This finding suggested the potential of terrapins as seed dispersers in eelgrass beds, which we sampled for two additional years (2010 and 2011). Seeds were only found in feces of terrapins captured prior to June 8 in all three years, coinciding with eelgrass seed maturation and release. Numbers of seeds in terrapin feces varied annually and decreased greatly in 2011 after an eelgrass die off in late 2010. The condition of seeds in terrapin feces was viable-mature, germinated, damaged, or immature. Of terrapins captured during time of seed release, 97% were males and juvenile females, both of which had head widths <30 mm. The fraction of individuals with ingested seeds was 33% for males, 35% for small females, and only 6% for large (mature) females. Probability of seed ingestion decreased exponentially with increasing terrapin head width; only males and small females (head width <30 mm) were likely to be vectors of seed dispersal. The characteristic that diamondback terrapins have well-defined home ranges allowed us to estimate the number of terrapins potentially dispersing eelgrass seeds annually. In seagrass beds of the Goodwin Islands region (lower York River, Virginia), there were 559 to 799 terrapins, which could disperse between 1,341 and 1,677 eelgrass seeds annually. These would represent a small proportion of total seed production within a single seagrass bed. However, based on probable home range distances, terrapins can easily traverse eelgrass meadow boundaries, thereby dispersing seeds beyond the bed of origin. Given the relatively short dispersion distance of eelgrass seeds, the diamondback terrapin may be a major source of inter-bed seed dispersal and genetic diversity.     

Regional differentiation among populations of the Diamondback terrapin (Malaclemys terrapin)

The Diamondback terrapin (Malaclemys terrapin) is a brackish-water turtle species whose populations have been fragmented due to anthropogenic activity such as development of coastal habitat and entrapment in commercial blue crab (Callinectes sapidus) fishing gear. Genetic analyses can improve conservation efforts for the long-term protection of the species. We used microsatellite DNA analysis to investigate levels of gene flow among and genetic variability within 21 geographically separate collections of the species distributed from Massachusetts to Texas. Quantified levels of genetic variability (allelic diversity, genotypic frequencies, and heterozygosity) revealed three zones of genetic discontinuity, resulting in four discrete populations: Northeast Atlantic, Coastal Mid-Atlantic, Florida and Texas/Louisiana. The average number of alleles and expected heterozygosity for the four genetic clusters were N_A = 6.54 and H_E = 0.050, respectively. However, the geographic boundaries of the populations did not correspond to accepted terrapin subspecies limits. Our results illuminate not only the need to sample terrapins in additional sites, specifically in the southeast, but also the necessity for allowing uninterrupted gene flow among population groupings to preserve current levels of genetic diversity.     

Geographic Variation in Egg Size and Lipid Provisioning in the Diamondback Terrapin Malaclemys terrapin

Abstract Understanding phenotypic differentiation among populations of wide-ranging species remains at the core of life-history research, because adaptation to local environmental conditions is expected. For example, when energy resources influence offspring fitness (as in oviparous ectotherms), the egg and hatchling environments are expected to influence selection by acting on the amount of energy allocated to offspring. Here we identify population variation in egg mass, length, width, and volume from diamondback terrapin Malaclemys terrapin eggs collected in Rhode Island (RI), Maryland (MD), and South Carolina (SC). Egg size (mean volume: 7.6, 8.1, and 9.1 cc in RI, MD, and SC, respectively) and clutch size (mean no. eggs: 16.1, 12.2, and 6.0 in RI, MD, and SC, respectively) differed among populations, which indicated that females produce larger clutches with smaller eggs at high latitudes and smaller clutches of larger eggs at lower latitudes. Lipid analyses indicated that eggs from SC contained yolks with a higher proportion of nonpolar lipids than did eggs from MD or RI (mean percentage of nonpolar lipids: 22.3%, 22.5%, and 31.8% in RI, MD, and SC, respectively). Thus, female terrapins in SC are laying larger eggs with increased lipid content to provide more energy for the developing embryo. Interestingly, total triacylglycerol (energetic lipid) was greater in southern populations but occurred in higher proportions in northern populations (total triacylglycerol: 88.0%, 85.4%, and 81.9% in RI, MD, and SC, respectively). This variation in triacylglycerol levels demonstrates the necessity for quantifying each lipid component. These data indicate a difference in reproductive strategy by which females in northern populations invest in higher fecundity with less energetic resources per offspring, whereas females in southern populations invest in larger eggs with considerably greater energy reserves.     

Spatiotemporal analysis of gene flow in Chesapeake Bay Diamondback Terrapins (Malaclemys terrapin)

There is widespread concern regarding the impacts of anthropogenic activities on connectivity among populations of plants and animals, and understanding how contemporary and historical processes shape metapopulation dynamics is crucial for setting appropriate conservation targets. We used genetic data to identify population clusters and quantify gene flow over historical and contemporary time frames in the Diamondback Terrapin (Malaclemys terrapin). This species has a long and complicated history with humans, including commercial overharvesting and subsequent translocation events during the early twentieth century. Today, terrapins face threats from habitat loss and mortality in fisheries bycatch. To evaluate population structure and gene flow among Diamondback Terrapin populations in the Chesapeake Bay region, we sampled 617 individuals from 15 localities and screened individuals at 12 polymorphic microsatellite loci. Our goals were to demarcate metapopulation structure, quantify genetic diversity, estimate effective population sizes, and document temporal changes in gene flow. We found that terrapins in the Chesapeake Bay region harbour high levels of genetic diversity and form four populations. Effective population sizes were variable. Among most population comparisons, estimates of historical and contemporary terrapin gene flow were generally low (m ≈ 0.01). However, we detected a substantial increase in contemporary gene flow into Chesapeake Bay from populations outside the bay, as well as between two populations within Chesapeake Bay, possibly as a consequence of translocations during the early twentieth century. Our study shows that inferences across multiple time scales are needed to evaluate population connectivity, especially as recent changes may identify threats to population persistence.     

Population genetics of the diamondback terrapin (Malaclemys terrapin)

We examined the population genetic structure of the diamondback terrapins (Malaclemys terrapin), within and among estuaries. Based on mark-recapture studies, these estuarine turtles have high site fidelity that is likely to make them vulnerable to local extinctions. We tested if observed site fidelity of adults would be reflected in intraestuarine population genetic structure of six highly polymorphic microsatellite loci (five tetranucleotide and one dinucleotide). No evidence was found for population structuring within the Charleston estuary nor among three different estuaries in South Carolina. We then examined four other terrapin populations from North Carolina to New York, as well as from the Florida Keys and from Texas. With increasing geographical distance, genetic differentiation increased from South Carolina through New York, but overall values were low. The dinucleotide locus contributed significantly more to the genetic differentiation of some population comparisons than any of the other loci. Interestingly, terrapins from South Carolina to New York were much more genetically similar to those from Texas (rho = 0.154) than to those from Florida (rho = 0.357). We attribute this pattern to extensive translocations of terrapins during the early 20th century to replenish diminished populations and to provide turtle farms with stocks. Terrapins collected in Texas were especially sought for shipment to the northeastern US because of their larger size. Our study indicates no population structure within or among adjacent estuaries. Thus, the mark-recapture information from adult and subadult feeding locations is a poor predictor of population genetic structure. Additionally, it appears that past human activities may have drastically altered the genetics of current populations. Finally, our data suggest that translocation of eggs or head starting of terrapins within estuaries or among adjacent estuaries is acceptable from a genetic standpoint.     

Purification and partial characterization of high and low activity carbonic anhydrase isoenzymes from Malaclemys terrapin centrata.

1. High activity (CA C) and low activity (CA B) carbonic anhydrase isoenzymes have been purified from turtle erythrocytes. 2. The two isoenzymes differed in CO2 hydration specific activity by 36-fold. 3. The low activity isoenzyme contained one half-cystine residue, whereas the high activity isoenzyme contained four half-cystines and required a reducing environment to maintain activity. Both isoenzymes contained zinc. 4. Molecular weights of 28,500 and 30,400 daltons were established for the low and high activity isoenzymes respectively. 5. Both isoenzymes were inhibited by acetazolamide, but only the high activity isoenzyme was inhibited by parachloromercuribenzoate. 6. The low activity isoenzyme was present in the erythrocytes at about 8-10 times the concentration of the high activity isoenzyme. 7. The high activity isoenzyme cross-reacted with antibodies prepared against pure chicken carbonic anhydrase C.     

Seasonal changes in thermal environment and metabolic enzyme activity in the diamondback terrapin (Malaclemys terrapin)

Diamondback terrapins experience broad fluctuations in temperature on both a daily and seasonal basis in their estuarine environment. We measured metabolic enzyme activity in terrapin muscle tissue to assess thermal dependence and the role of temperature in seasonal metabolic downregulation in this species. Activity of lactate dehydrogenase (LDH), pyruvate kinase (PK), citrate synthase (CS), and cytochrome c oxidase (CCO) was assayed at 10, 20, 30, and 40 °C for tissue collected during summer and winter. The Q(10) for enzyme activity varied between 1.31 and 2.11 within the temperature range at which terrapins were active (20-40 °C). The Q(10) for LDH, CS, and CCO varied between 1.39 and 1.76 and between 10 and 20 °C, but PK exhibited heightened thermal sensitivity within this lower temperature range, with a Q(10) of 2.90 for summer-collected tissue and 5.55 for winter-collected tissue. There was no significant effect of season on activity of LDH or PK, but activity of CS and CCO was significantly lower in winter-collected tissue compared with summer-collected tissue. Results indicate that temperature effects contribute to seasonal metabolic downregulation and dormancy in terrapins, but other environmental factors (i.e. oxygen availability), as well as seasonal shifts in blood biochemistry and circulating hormones may also play an important role.     

Seasonal Enumeration of Fecal Coliform Bacteria from the Feces of Ring-Billed Gulls (Larus delawarensis) and Canada Geese (Branta canadensis)

Water suppliers have often implicated roosting birds for fecal contamination of their surface waters. Geese and gulls have been the primary targets of this blame although literature documenting the fecal coliform content of these birds is quite limited. To determine the actual fecal coliform concentrations of these birds, fecal samples from 249 ring-billed gulls and 236 Canada geese in Westchester County, N.Y., were analyzed over a 2-year period. Results indicate that gull feces contain a greater average concentration of fecal coliform bacteria per gram (3.68 × 10⁸) than do goose feces (1.53 × 10⁴); however, average fecal sample weights of the geese were more than 15 times higher than those of the gulls.     

Maximum standard metabolic rate corresponds with the salinity of maximum growth in hatchlings of the estuarine northern diamondback terrapin (Malaclemys terrapin terrapin): Implications for habitat conservation

I evaluated standard metabolic rates (SMR) of hatchling northern diamondback terrapins (Malaclemys terrapin terrapin) across a range of salinities (salinity = 1.5, 4, 8, 12, and 16 psu) that they may encounter in brackish habitats such as those in the Maryland portion of the Chesapeake Bay, U.S.A. Consumption of O₂ and production of CO₂ by resting, unfed animals served as estimates of SMR. A peak in SMR occurred at 8 psu which corresponds closely with the salinity at which hatchling growth was previously shown to be maximized (salinity ∼ 9 psu). It appears that SMR is influenced by growth, perhaps reflecting investments in catabolic pathways that fuel anabolism. This ecophysiological information can inform environmental conservation and management activities by identifying portions of the estuary that are bioenergetically optimal for growth of hatchling terrapins. I suggest that conservation and restoration efforts to protect terrapin populations in oligo-to mesohaline habitats should prioritize protection or creation of habitats in regions where average salinity is near 8 psu and energetic investments in growth appear to be maximized.     

Comparison of Gelman and Millipore Membrane Filters for Enumerating Fecal Coliform Bacteria

Tests of two leading brands of membrane filters used for enumerating fecal coliform bacteria showed that Gelman GN-6 filters recovered statistically more colonies of bacteria than did Millipore HAWG 047SO filters from pure cultures incubated at either 35 C (the optimal growth temperature) or 44.5 C (the standard temperature for the fecal coliform test). Standard membrane filter procedures with M-FC broth base were used to enumerate the organisms. Densities of colonies incubated on Gelman filters at 44.5 C averaged 2.3 times greater than those on Millipore filters. Plate counts of the bacteria at both temperatures indicated that incubation at 44.5 C did not inhibit propagation of fecal coliform bacteria. For the pour plates, M-FC broth base plus 1.5% agar was used. This modified medium compared favorably to plate count agar for enumerating Escherichia coli. At 35 and 44.5 C, colony counts on Gelman filters agreed closely with plate counts prepared concurrently, but Millipore counts were consistently lower than plate counts, especially at 44.5 C. Comparative analyses of river water for fecal coliform bacteria by the membrane filter technique gave results comparable to those for the pure cultures.     

Relationships of maternal body size and morphology with egg and clutch size in the diamondback terrapin,Malaclemys terrapin (Testudines: Emydidae)

Because resources are finite, female animals face trade‐offs between the size and number of offspring they are able to produce during a single reproductive event. Optimal egg size (OES) theory predicts that any increase in resources allocated to reproduction should increase clutch size with minimal effects on egg size. Variations of OES predict that egg size should be optimized, although not necessarily constant across a population, because optimality is contingent on maternal phenotypes, such as body size and morphology, and recent environmental conditions. We examined the relationships among body size variables (pelvic aperture width, caudal gap height, and plastron length), clutch size, and egg width of diamondback terrapins from separate but proximate populations at Kiawah Island and Edisto Island, South Carolina. We found that terrapins do not meet some of the predictions of OES theory. Both populations exhibited greater variation in egg size among clutches than within, suggesting an absence of optimization except as it may relate to phenotype/habitat matching. We found that egg size appeared to be constrained by more than just pelvic aperture width in Kiawah terrapins but not in the Edisto population. Terrapins at Edisto appeared to exhibit osteokinesis in the caudal region of their shells, which may aid in the oviposition of large eggs.