Characterising injuries and pathologies of common dolphin Delphinus delphis mortalities in the South Australian Sardine Fishery.

Mortality of dolphins in fishing operations is often under-estimated, as shown by studies of beach-washed carcasses. Linking evidence obtained during necropsies with fishing method is fundamental to understanding the extent of mortality and the manner in which animals die. The South Australian Sardine Fishery (SASF) has operated a purse-seine industry since 1991. This study characterised injuries, pathological changes and life history of 49 dead dolphins collected from SASF during 2006-2019. Histology examination was conducted on 25 animals. Neonates, calves and juveniles accounted for 63% of the sample. Of mature females (n = 14), 11 were pregnant or lactating, with cryptic mortality estimated to be 20% of dolphins studied. Body condition was robust in 48 dolphins. Net marks were seen on 82%, mostly on the head, trunk and peduncle. Broken/missing teeth were noted in 63%. All dolphins had subdermal haemorrhage (moderate to severe in 96%), particularly around the head. Deep haemorrhage was common, including around occipital and flipper condyles, and organs. Copious fluid was present in the thoracic (pleural) and abdominal (ascites) cavities of half of the dolphins. Within the lungs, watery fluid and froth were observed in 100 and 39%, respectively. Recent bone fractures were documented in 43% of dolphins, mostly associated with haemorrhage. Severe blunt trauma appeared to be the primary cause of death, and 10 dolphins also had other significant pathologies. Visceral organ congestion and mild cardiomyopathy were observed. Stomachs contained prey remains in 75% of cases. The results of this study may help identify unreported purse-seine mortalities washed up in South Australia and elsewhere.

A review of heavy metal and organochlorine levels in marine mammals in Australia.

Study of toxic contaminants in marine mammal specimens collected around Australia is currently uncoordinated and piecemeal. Most states collect samples but there is little or no financial support for their analysis. This study combines data, published or unpublished, from 13 sources. Heavy metals have been analysed in about 676 specimens; over 400 were for mercury levels in P. macrocephalus taken at a whaling station. The remaining samples were mostly from toothed whales, a few baleen whales (< 20), pinnipeds (41) and dugongs (49). The most consistently analysed metals were lead, mercury and cadmium. Liver and kidney lead levels ranged from < 1-3 ppm; levels in bone were 0-418 ppm, with most less than 10 ppm. Mercury levels in a large sample of P. macrocephalus muscle were < 12.2 ppm. Mercury levels in the small number of samples from other species were 0.51-143 ppm (kidney), 1.52-479 ppm (liver) and < 0.1-36 ppm (muscle). Cadmium levels in liver (0-52 ppm) and kidney (0-106 ppm) were extremely variable. Levels greater than 10 ppm were recorded in many species and were especially high in Hydrurga leptonyx, Dugong dugon, Mesoplodon layardii and Pseudorca crassidens. Adult Tursiops truncatus inhabiting the inshore gulfs of South Australia had considerably higher levels of cadmium compared with other regions. Information on organochlorine levels is sparse (approximately 39 specimens) and suggest low levels when compared to other parts of the world. Total DDT was highest (28.4 ppm) in a neonatal Orcinus orca. Some high levels of DDT were recorded in Tursiops truncatus, Delphinus delphis and Arctocephalus pusillus doriferus. PCBs ranged from < 0.05 to 3.87 ppm. A comprehensive pathological assessment of marine mammals is needed in order to evaluate the effects of toxic contaminants.

An improved, luminescent europium-based stain for detection of electroblotted proteins on nitrocellulose or polyvinylidene difluoride membranes.

SYPRO Rose Plus protein blot stain is an improved europium-based metal chelate stain for the detection of proteins on nitrocellulose and poly(vinylidene difluoride) (PVDF) membranes. Staining is achieved without covalently modifying the proteins. The stain may be excited with a 254 nm (UV-C), 302 nm (UV-B), or 365 nm (UV-A) light source and displays a sharp emission maximum at 612 nm. The emission peak has a full width at half-maximum of only 8 nm. The stain exhibits exceptional photostability, allowing long exposure times for maximum sensitivity. Since the dye is composed of a europium complex, it has a long emission lifetime, potentially allowing time-resolved detection, greatly reducing background fluorescence. Proteins immobilized to a nitrocellulose or PVDF membrane by electroblotting, dot-blotting, or vacuum slot-blotting are incubated with SYPRO Rose Plus protein blot stain for 15-30 min. Membranes are rinsed briefly, visualized with UV epi-illumination and the luminescence of the europium dye is measured using a 490 nm long-pass or 625 +/- 15 nm band-pass filter in combination with a conventional photographic or charge-coupled device (CCD) camera system. Alternatively, the dye may be visualized using a xenon-arc illumination source. The stain is readily removed from proteins by incubating membranes at mildly alkaline pH. The reversibility of the protein staining procedure allows for subsequent biochemical analyses, such as immunoblotting and biotin-streptavidin detection using colorimetric, direct fluorescence or fluorogenic visualization methods.

Green/red dual fluorescence detection of total protein and alkaline phosphate-conjugated probes on blotting membranes.

A two-color fluorescence detection method is described based upon covalently coupling the succinimidyl ester of BODIPY FL-X to proteins immobilized on poly(vinylidene difluoride) (PVDF) membranes, followed by detection of target proteins using the fluorogenic substrate 9H-(1,3-dichloro-9,9-dimethylacridin-2-one-7-yl(DDAO)-phosphate in combination with alkaline-phosphatase-conjugated reporter molecules. This results in all proteins in the profile being visualized as green signal while those detected specifically with the alkaline-phosphatase conjugate appear as red signal. The dichromatic detection system is broadly compatible with a wide range of analytical imaging devices including UV epi- or transilluminators combined with photographic or charge-coupled device (CCD) cameras, xenon-arc sources equipped with appropriate excitation/emission filters, and dual laser gel scanners outfitted with a 473 nm second-harmonic generation or 488 nm argon-ion laser as well as a 633 nm helium-neon or 635 nm diode laser. The dichromatic detection method permits detection of low nanogram amounts of protein and allows for unambiguous identification of target proteins relative to the entire protein profile on a single electroblot, obviating the need to run replicate gels that would otherwise require visualization of total proteins by silver staining and subsequent alignment with chemiluminescent or colorimetric signals generated on electroblots.