Comparison of the marine soundscape before and during the COVID-19 pandemic in dolphin habitat in Sarasota Bay, FL.

During the COVID-19 pandemic, changes in vessel activity and associated noise have been reported globally. Sarasota Bay is home to a large and increasing number of recreational vessels as well as a long-term resident community of bottlenose dolphins, Tursiops truncatus. Data were analyzed from two hydrophones to compare the soundscape during the COVID-19 pandemic to previous years (March-May 2020 and 2018/2019). Hourly metrics were calculated: vessel passes, 95th percentile sound levels [125 Hz and 16 kHz third octave bands (TOBs), and two broader bands: 88-1122 Hz and 1781-17 959 Hz], and dolphin whistle detection to understand changes in vessel activity and the effect on wildlife. Vessel activity increased during COVID-19 restrictions by almost 80% at one site and remained the same at the other site. Of the four sound level measures, only the 125 Hz TOB and 88-1122 Hz band increased with vessel activity at both sites, suggesting that these may be appropriate measures of noise from rapid pass-bys of small vessels in very shallow (<10 m) habitats. Dolphin whistle detection decreased during COVID-19 restrictions at one site but remained the same at the site that experienced increased vessel activity. The results suggest that pandemic effects on wildlife should not be viewed as homogeneous globally.

Y chromosome sequence variation and the history of human populations.

Binary polymorphisms associated with the non-recombining region of the human Y chromosome (NRY) preserve the paternal genetic legacy of our species that has persisted to the present, permitting inference of human evolution, population affinity and demographic history. We used denaturing high-performance liquid chromatography (DHPLC; ref. 2) to identify 160 of the 166 bi-allelic and 1 tri-allelic site that formed a parsimonious genealogy of 116 haplotypes, several of which display distinct population affinities based on the analysis of 1062 globally representative individuals. A minority of contemporary East Africans and Khoisan represent the descendants of the most ancestral patrilineages of anatomically modern humans that left Africa between 35,000 and 89,000 years ago.

Bubbles in live-stranded dolphins.

Bubbles in supersaturated tissues and blood occur in beaked whales stranded near sonar exercises, and post-mortem in dolphins bycaught at depth and then hauled to the surface. To evaluate live dolphins for bubbles, liver, kidneys, eyes and blubber-muscle interface of live-stranded and capture-release dolphins were scanned with B-mode ultrasound. Gas was identified in kidneys of 21 of 22 live-stranded dolphins and in the hepatic portal vasculature of 2 of 22. Nine then died or were euthanized and bubble presence corroborated by computer tomography and necropsy, 13 were released of which all but two did not re-strand. Bubbles were not detected in 20 live wild dolphins examined during health assessments in shallow water. Off-gassing of supersaturated blood and tissues was the most probable origin for the gas bubbles. In contrast to marine mammals repeatedly diving in the wild, stranded animals are unable to recompress by diving, and thus may retain bubbles. Since the majority of beached dolphins released did not re-strand it also suggests that minor bubble formation is tolerated and will not lead to clinically significant decompression sickness.

Demographic Assessment of Mono(2-ethylhexyl) Phthalate (MEHP) and Monoethyl Phthalate (MEP) Concentrations in Common Bottlenose Dolphins (Tursiops truncatus) From Sarasota Bay, FL, USA.

Common bottlenose dolphins (Tursiops truncatus) have previously demonstrated exposure to phthalate esters. Phthalates and phthalate esters are commonly added to consumer goods to enhance desirable properties. As the amount of plastic marine debris increases, these chemicals can easily leach from these products into the surrounding environment. To evaluate demographic variability in exposure, eight phthalate metabolites were quantified in urine samples collected from free-ranging bottlenose dolphins sampled in Sarasota Bay, FL, USA (2010-2019; n = 51). Approximately 75% of individual dolphins had detectable concentrations of at least one phthalate metabolite. The most frequently detected metabolites were mono(2-ethylhexyl) phthalate (MEHP; n = 28; GM = 4.57 ng/mL; 95% CI = 2.37-8.80; KM mean = 7.95; s.d. = 15.88) and monoethyl phthalate (MEP; GM = 4.51 ng/mL; 95% CI = 2.77-7.34; ROS mean = 2.24; s.d. = 5.58). Urinary concentrations of MEHP and MEP were not significantly different between sex (MEHP p = 0.09; MEP p = 0.22) or age class (i.e., calf/juvenile vs. adult; MEHP p = 0.67; MEP p = 0.13). Additionally, there were no significant group differences in the likelihood of MEHP or MEP detection for any demographic as determined by a Peto-Peto test. Frequency of detection was similar for both metabolites between males and females (MEHP p = 0.10; MEP p = 0.40) as well as between juveniles and adults (MEHP p = 0.50; MEP: p = 0.60). These findings suggest ubiquitous exposure risk for both sexes and age classes, warranting further investigation into potential sources and health implications.

African origin of modern humans in East Asia: a tale of 12,000 Y chromosomes.

To test the hypotheses of modern human origin in East Asia, we sampled 12,127 male individuals from 163 populations and typed for three Y chromosome biallelic markers (YAP, M89, and M130). All the individuals carried a mutation at one of the three sites. These three mutations (YAP+, M89T, and M130T) coalesce to another mutation (M168T), which originated in Africa about 35,000 to 89,000 years ago. Therefore, the data do not support even a minimal in situ hominid contribution in the origin of anatomically modern humans in East Asia.

Prevalence and pathology of lungworm infection in bottlenose dolphins Tursiops truncatus from southwest Florida.

Parasitism of the respiratory system is a relatively common finding in stranded cetaceans; however, no systematic investigations regarding the severity, distribution, and clinical consequences of these infections in bottlenose dolphins Tursiops truncatus have been conducted previously. The present study determined the prevalence of lungworm infections in dead stranded (n=22) and live bottlenose dolphins (n=44) from southwestern Florida, USA, during the period from 2003 to 2005. Dead stranded bottlenose dolphins were necropsied and lungs were examined visually, by palpation, and histologically for lesions consistent with verminous pneumonia. When present, nematodes were counted, measured, and identified to species based upon their morphology. Dolphin feces and blowhole swabs were collected and examined for nematode larvae. Lungworm prevalence was 77% in dead animals (n=22). The lesions in most cases were mild, chronic, and not the primary cause of death. Only 13% of dead animals examined had patent infections, with larvae present in blowhole and fecal cytology, and only 18% of animals had intact worms present at necropsy, with a geometric mean intensity of infection of 22.6 worms animal(-1). Intact worms were identified as either Halocercus lagenorhynchi or Skrjabinalius cryptocephalus. The highest prevalence of active infections was found in neonates and calves, including 1 stillborn calf. For free-ranging animals, all blowhole swabs (n=44) were negative, and fecal cytology (n=22) showed a 3% prevalence of patent infection. Findings from the present study support the theory that bottlenose dolphins can be infected transplacentally by lungworms. The impact that such infections may have on neonatal survival is unknown; however, these infections could increase neonatal mortality.

Dietary cation-anion difference may explain why ammonium urate nephrolithiasis occurs more frequently in common bottlenose dolphins () under human care than in free-ranging common bottlenose dolphins.

Ammonium urate nephrolithiasis frequently develops in common bottlenose dolphins () managed under human care but is rare in free-ranging common bottlenose dolphins. In other species, the dietary cation-anion difference (DCAD) can affect ammonium urate urolith formation by increasing proton excretion as ammonium ions. Therefore, differences in diet between the 2 dolphin populations could affect urolith formation, but the DCAD of most species consumed by free-ranging and managed dolphins is unknown. To compare the nutrient composition of diets consumed by free-ranging and managed bottlenose dolphins, samples ( = 5) of the 8 species of fish commonly consumed by free-ranging bottlenose dolphins in Sarasota Bay, FL, and the 7 species of fish and squid commonly fed to managed bottlenose dolphins were analyzed for nutrient content. Metabolizable energy was calculated using Atwater factors; the DCAD was calculated using 4 equations commonly used in people and animals that use different absorption coefficients. The nutrient composition of individual species was used to predict the DCAD of 2 model diets typically fed to managed common bottlenose dolphins and a model diet typically consumed by common bottlenose dolphins in Sarasota Bay. To mimic differences in postmortem handling of fish for the 2 populations of bottlenose dolphins, "free-ranging" samples were immediately frozen at -80°C and minimally thawed before analysis, whereas "managed" samples were frozen for 6 to 9 mo at -18°C and completely thawed. "Free-ranging" species contained more Ca and P and less Na and Cl than "managed" fish and squid species. As a consequence, the DCAD of both model managed dolphin diets obtained using 3 of the 4 equations was much more negative than the DCAD of the model free-ranging bottlenose dolphin diet ( < 0.05). The results imply that managed bottlenose dolphins must excrete more protons in urine than free-ranging bottlenose dolphins, which will promote nephrolith formation. The nutrient composition of the free-ranging bottlenose dolphin diet, determined for the first time here, can be used as a guide for feeding managed bottlenose dolphins, but research in vivo is warranted to determine whether adding more cations to the diet will prevent urolith formation in managed dolphins.

Role of anchorage in the expression of tumorigenicity of untransformed mouse cell lines.

Cultured mouse cells were tested for tumorigenicity in nude mice with both a conventional assay (injection of cell suspensions) and a new test involving implantation of cells grown on gelatin sponges. Sublines of Balb/3T3 cells, obtained from different sources, varied in their tumorigenic potential with either assay. One subline (A) formed distinctive precancerous nodules only in the sponge assay; these nodules often became progressive after a latent period of 3-4 months. However, suspensions of cells of this subline also caused tumors after a similar latent period, but no nodular phase preceded tumor formation. Another subline of Balb/3T3 (M) has failed to form tumors in either assay. The Balb/3T3 sublines did not differ in vitro properties, such as low saturation density, failure to grow in methylcellulose, and monolayer morphology. A second experimental approach involved tests on nude BALB/c mouse-embryo fibroblasts at various passage levels. The cells were passaged from primary culture, through crisis, to heteroploid, established cell lines. Tumorigenicity was demonstrable earlier in the sponge assay, at which time in vitro parameters putatively associated with malignant behavior were unchanged. Possible relationships with the in vivo phenomenon of solid-surface sarcomagenesis are discussed.

An in vitro assay for growth regulation of embryonal carcinoma by the blastocyst.

An in vitro assay has been developed in order to examine the mechanism whereby the mouse blastocyst regulates embryonal carcinoma (EC). The assay measures the suppression of EC cell colony formation caused by exposure to the cavity of the blastocyst (i.e., blastocele), and the results are comparable to the previous results with blastocyst-mediated suppression of tumor formation for this same cell line. The assay has been used to determine the time necessary for the blastocyst to regulate the EC cell. In these experiments, immunosurgery is done to disrupt the interaction between the EC cell and the blastocyst, and the EC cell is then recovered and identified by its ability to form a colony. As compared to control cells, 84% of the EC cells are recovered after 2 hr of exposure to the blastocyst, 57% are recovered after 14 hr of exposure, and 27% are recovered after 24 hr of exposure. This long time course is similar to the prolonged doubling time of labeled EC cells within the blastocyst suggesting that the response of the EC cell may be related to the cell cycle within the blastocyst. Data presented show that synchronization of the EC cell cycle (using a mitotic selection procedure) produces synchronization of the response to the blastocyst. The response appears to be closely linked to the G1 phase of the EC cell cycle.

Widespread inhibition of neuroblastoma cells in the 13- to 17-day-old mouse embryo.

Previous experiments have demonstrated specific inhibition of tumor formation after neuroblastoma cells were injected into fragments of 8.5- to 9.5-day embryonic tissue (A.H. Podesta et al. Proc. Natl. Acad. Sci. USA, 81:7608-7611, 1984). The effect was localized to the somite and appeared specific for neuroblastoma as opposed to a variety of other tumor types. This regulation of neuroblastoma cells was believed to reflect an underlying event in the development of migrating embryonic neuroblasts. The current experiments were done to determine the effect on regulation with further embryonic development. The results indicated that later embryos (13 to 17 days of gestation) have a widespread inhibitory effect in all tissues tested, including the adrenal gland, testis, kidney, liver, limb bud, and heart. In contrast a leukemia cell line was not affected by any of these tissues. In organ culture demonstrable colony formation by neuroblastoma was likewise inhibited, and conditioned media from one of these embryonic sources (limb bud) slowed but did not abrogate growth of neuroblastoma cells.

Specificity of the control of tumor formation by the blastocyst.

An assay to determine the mechanism of regulation of embryonal carcinoma cells by the blastocyst, which is based on a comparison of tumors produced when the cancer cells are cloned alone or after incorporation into blastocysts, was refined by labeling embryonal carcinoma cells with fluorescent microspheres and by following their fate after injection into the blastocysts. Through the use of the new techniques, it was observed that cells of one line of nullipotent embryonal carcinoma were controlled at the 50% level, those from another were not controlled, and those from a multipotent but undifferentiated line were controlled in almost absolute fashion. Single Sarcoma 180 of L1210 leukemia cells were not controlled when injected into the blastocele, but C1300 neuroblastoma cells were partially controlled. None of these tumors have a normal cellular counterpart in the blastocyst, as does embryonal carcinoma, but neurulation follows blastulation by only a few days, so that the neuroblastoma cells may be regulated at that time. Parietal yolk sac carcinoma cells, which have a counterpart in the late blastocyst, were not controlled. On the basis of these data, it is postulated that, if one embryonic field can regulate its closely related cancer, then there may be an embryonic field capable of regulating each carcinoma.

Trophectoderm in control of murine embryonal carcinoma.

It has been shown previously that the intact blastocyst of the mouse can regulate tumor formation and colony formation of murine embryonal carcinoma. This effect is consistent with the close histogenetic correspondence between embryonal carcinoma and the inner cell mass of the blastocyst. The ability of inner cell mass, blastocele fluid, and inner and outer surfaces of trophectoderm to abrogate colony formation of a variety of malignant tumors has now been tested. Direct contact of the embryonal carcinoma cells with the blastocele surface of trophectoderm proved to be necessary for abrogation of colony formation of embryonal carcinoma. This effect was not seen with any of the other tumors tested. Some tumors, which lack a normal cellular counterpart in the blastocyst, grew poorly in the blastocele unless a fistula was made in the wall of the blastocyst. Colony formation of the embryonal carcinoma was regulated in blastocysts with fistulas, but the other tumors were not regulated under these conditions. It is concluded that colony formation of embryonal carcinoma cells is regulated by direct contact with the trophectoderm of its corresponding embryonic field in an unknown but specific manner.

A Targeted Metabolomics Assay to Measure Eight Purines in the Diet of Common Bottlenose Dolphins, Tursiops truncatus.

Bottlenose dolphins managed under human care, human beings and Dalmatian dogs are prone to forming urate uroliths. Limiting dietary purine intake limits urate urolith formation in people and dogs because purines are metabolized to uric acid, which is excreted in urine. Managed dolphins develop ammonium urate nephroliths, whereas free-ranging dolphins do not. Free-ranging dolphins consume live fish, whereas managed dolphins consume different species that have been stored frozen and thawed. Differences in the purine content of fish consumed by dolphins under human care versus in the wild may be responsible for the difference in urolith prevalence. Commercially available purine assays measure only four purines, but reported changes in purines during frozen storage suggest that a wider range of metabolites should be measured when comparing fresh and stored fish. A method using high performance liquid chromatography with tandem mass spectrometry was developed to quantify eight purine metabolites in whole fish and squid commonly consumed by dolphins. The coefficient of variation within and among days was sometimes high for purines present in small amounts but was acceptable (≤ 25%) for guanine, hypoxanthine, and inosine, which were present in high concentrations. This expanded assay identified a total purine content up to 2.5 times greater than the total that would be quantified if only four purines were measured. Assuming additional purines are absorbed, these results suggest that additional purine metabolites should be measured to better understand the associated risk when fish or other purine-rich foods are consumed by people or animals prone to developing uroliths.

Nucleotide variation at the Gpdh locus in the genus Drosophila.

The Gpdh locus was sequenced in a broad range of Drosophila species. In contrast to the extreme evolutionary constraint seen at the amino acid level, the synonymous sites evolve at rates comparable to those of other genes. Gpdh nucleotide sequences were used to infer a phylogenetic tree, and the relationships among the species of the obscura group were examined in detail. A survey of nucleotide polymorphism within D. pseudoobscura revealed no amino acid variation in this species. Applying a modified McDonald-Kreitman test, the amino acid divergence between species in the obscura group does not appear to be excessive, implying that drift is adequate to explain the patterns of amino acid change at this locus. In addition, the level of polymorphism at the Gpdh locus in D. pseudoobscura is comparable to that found at other loci, as determined by a Hudson-Kreitman-Aguadé test. Thus, the pattern of nucleotide variation within and between species at the Gpdh locus is consistent with a neutral model.

Genetics and dermatology: a review.

The development of genetic studies over the last hundred years, in particular as they have affected dermatology, has been reviewed. The clinical application of knowledge derived from other fields has been emphasized, areas of future research have been indicated, and suggestions for cooperation with other disciplines have been offered.

The histology of ichthyosis.

Histological features of 12 biopsy specimens from patients with ichthyosis inherited as a dominant trait were compared with 15 specimens from patients with ichthyosis inherited as a sex-linked recessive trait and with 12 control sections from normal skin. Characteristic features that distinguished each inherited variety were described. The histology of sex-linked ichthyosis is very similar to that of non-bullous ichthyosiform erythrodermia.

Excessive homoplasy in an evolutionarily constrained protein.

The evolution of monomorphic proteins among closely related species has not been examined in detail. To investigate this phenomenon, the glycerol-3-phosphate dehydrogenase (Gpdh) locus was sequence in a broad range of Drosophila species. Although purifying selection to remove amino acid variation is the dominant force in the evolution of Gpdh, some replacements have occurred. The sequences were compared in the context of the phylogeny of the genus, revealing a high proportion of amino acid parallelism and reversal (homoplasy) at four sites. The level of homoplasy is significantly greater than that seen in other proteins for which multiple sequences are available, showing that Gpdh is strongly constrained by both the number of amino acid differences and the types of changes allowed. These four sites evolve at a much higher rate than do the other variable positions in the protein, accounting for half of the interspecific amino acid replacements. However, unlike typical hypervariable sites, where multiple changes to several different amino acids are seen, evolutionary 'flip-flopping' between two amino acid states defines this new class of hypervariable site.

Epidermal diseases in bottlenose dolphins: impacts of natural and anthropogenic factors.

Experimental studies have highlighted the potential influence of contaminants on marine mammal immune function and anthropogenic contaminants are commonly believed to influence the development of diseases observed in the wild. However, estimates of the impact of contaminants on wild populations are constrained by uncertainty over natural variation in disease patterns under different environmental conditions. We used photographic techniques to compare levels of epidermal disease in ten coastal populations of bottlenose dolphins (Tursiops truncatus) exposed to a wide range of natural and anthropogenic conditions. Epidermal lesions were common in all populations (affecting > 60% of individuals), but both the prevalence and severity of 15 lesion categories varied between populations. No relationships were found between epidermal disease and contaminant levels across the four populations for which toxicological data were available. In contrast, there were highly significant linear relationships with oceanographic variables. In particular, populations from areas of low water temperature and low salinity exhibited higher lesion prevalence and severity. Such conditions may impact on epidermal integrity or produce more general physiological stress, potentially making animals more vulnerable to natural infections or anthropogenic factors. These results show that variations in natural environmental factors must be accounted for when investigating the importance of anthropogenic impacts on disease in wild marine mammals.

Decreased lymphocyte responses in free-ranging bottlenose dolphins (Tursiops truncatus) are associated with increased concentrations of PCBs and DDT in peripheral blood.

Since 1987, large-scale mortalities of dolphins have been reported along the Atlantic coast of North America, in the Gulf of Mexico, and in the Mediterranean Sea. Autopsied bottlenose dolphins, Tursiops truncatus, which were collected from the large-scale mortality along the Atlantic coast in 1987 to 1988, exhibited opportunistic infections indicative of immune dysfunction. Further, these animals had high levels of chlorinated hydrocarbons, such as PCBs and DDT, that can suppress immune functions. The purpose of this study was to determine whether there is a relationship between chemical contaminant exposure and immune response in free-ranging dolphins. In June of 1991, peripheral blood was obtained from members of a bottlenose dolphin population that resides along the west coast of Florida. Peripheral blood lymphocyte responses to Concanavalin A (Con A) and phytohemagglutinin (PHA) were determined in vitro and compared by regression analysis with contaminant concentrations in whole blood from a small subset of these animals (n = 5). These data indicate that a reduced immune response in these bottlenose dolphins was correlated with increasing whole blood concentrations of several contaminants. Specifically, inverse correlations were found between Con A-induced lymphocyte proliferation and tetrachlorinated to octachlorinated biphenyls (r2 values ranged from 0.70 to 0.87). Con A-induced lymphocyte responses also correlated inversely with p,p'DDT (r2 values of 0.73 and 0.79); o.p'-DDE (r2 values of 0.93 and 0.96); and p,p'-DDE (r2 values of 0.73 and 0.81).

Biomarkers of DNA damage in marine mammals.

Certain environmental contaminants found in marine mammals have been shown to cause DNA damage and cancer. The micronuclei (MN), sister chromatid exchange (SCE) and/or chromosome aberration (CA) assays were used to assess baseline (spontaneous) levels of DNA damage in blood lymphocytes of individuals of the relatively healthy and lightly contaminated Arctic beluga whale (Delphinapterus leucas), Sarasota Bay, FL, bottlenose dolphin (Tursiops truncatus) and Northwestern Atlantic grey (Halichoerus grypus) and harp (Phoca groenlandicus) seal populations. MN cell (MNC) frequencies ranged between 2 and 14/1000 binucleated (BN) cells and were statistically similar between species. In bottlenose dolphins, MNC frequency was correlated with age and was significantly higher in females than in males. No intraspecific variation in MNC frequency was found in beluga whales. Intraspecific variation was not tested in seals due to the small sample size. Frequencies of SCEs and total CAs, excluding gaps, ranged, respectively, between 1 and 15 SCE(s)/per cell and 4-6 CAs/100 cells in beluga whales. SCE and CA frequencies did not vary with age or sex in beluga whales. The MN, SCE and CA assays were found to be practical tools for the detection of DNA damage in marine mammals and could be used in the future to compare DNA damage between relatively lightly and highly contaminated populations.