Humpback whale migrations to Antarctic summer foraging grounds through the southwest Pacific Ocean.

Humpback whale (Megaptera novaeangliae) populations typically undertake seasonal migrations, spending winters in low latitude breeding grounds and summers foraging in high latitude feeding grounds. Until recently, a broad scale understanding of whale movement has been derived from whaling records, Discovery marks, photo identification and genetic analyses. However, with advances in satellite tagging technology and concurrent development of analytical methodologies we can now detail finer scale humpback whale movement, infer behavioural context and examine how these animals interact with their physical environment. Here we describe the temporal and spatial characteristics of migration along the east Australian seaboard and into the Southern Ocean by 30 humpback whales satellite tagged over three consecutive austral summers. We characterise the putative Antarctic feeding grounds and identify supplemental foraging within temperate, migratory corridors. We demonstrate that Antarctic foraging habitat is associated with the marginal ice zone, with key predictors of inferred foraging behaviour including distance from the ice edge, ice melt rate and variability in ice concentration two months prior to arrival. We discuss the highly variable ice season within the putative foraging habitat and the implications that this and other environmental factors may have on the continued strong recovery of this humpback whale population.

Pseudogenes and DNA-based diet analyses: a cautionary tale from a relatively well sampled predator-prey system.

Mitochondrial ribosomal DNA is commonly used in DNA-based dietary analyses. In such studies, these sequences are generally assumed to be the only version present in DNA of the organism of interest. However, nuclear pseudogenes that display variable similarity to the mitochondrial versions are common in many taxa. The presence of nuclear pseudogenes that co-amplify with their mitochondrial paralogues can lead to several possible confounding interpretations when applied to estimating animal diet. Here, we investigate the occurrence of nuclear pseudogenes in fecal samples taken from bottlenose dolphins (Tursiops truncatus) that were assayed for prey DNA with a universal primer technique. We found pseudogenes in 13 of 15 samples and 1-5 pseudogene haplotypes per sample representing 5-100% of all amplicons produced. The proportion of amplicons that were pseudogenes and the diversity of prey DNA recovered per sample were highly variable and appear to be related to PCR cycling characteristics. This is a well-sampled system where we can reliably identify the putative pseudogenes and separate them from their mitochondrial paralogues using a number of recommended means. In many other cases, it would be virtually impossible to determine whether a putative prey sequence is actually a pseudogene derived from either the predator or prey DNA. The implications of this for DNA-based dietary studies, in general, are discussed.

Islands in the sea: extreme female natal site fidelity in the Australian sea lion, Neophoca cinerea.

Pinnipeds (seals, fur seals, sea lions and walrus) form large breeding aggregations with females often remaining faithful to a natal site or area. In these cases, females are philopatric to regional areas on broad geographical scales of hundreds to thousands of kilometers. An investigation of variation in a control region sequence of mtDNA in the Australian sea lion (Neophoca cinerea) has shown a case of extreme female natal site fidelity that has resulted in almost fixed population differentiation across its range (PhiST=0.93). This high level of population subdivision over short geographical distances (approx. 60 km) is unparalleled in any social marine mammal and reflects the unique life-history traits of this rare species. The high level of population subdivision and exclusive female natal site fidelity has important ramifications for conservation management, and poses many interesting questions of both academic and applied interest.

Molecular scatology as a tool to study diet: analysis of prey DNA in scats from captive Steller sea lions.

The DNA of prey present in animal scats may provide a valuable source of information for dietary studies. We conducted a captive feeding trial to test whether prey DNA could be reliably detected in scat samples from Steller sea lions (Eumetopias jubatus). Two sea lions were fed a diet of fish (five species) and squid (one species), and DNA was extracted from the soft component of collected scats. Most of the DNA obtained came from the predator, but prey DNA could be amplified using prey-specific primers. The four prey species fed in consistent daily proportions throughout the trial were detected in more than 90% of the scat DNA extractions. Squid and sockeye salmon, which were fed as a relatively small percentage of the daily diet, were detected as reliably as the more abundant diet items. Prey detection was erratic in scats collected when the daily diet was fed in two meals that differed in prey composition, suggesting that prey DNA is passed in meal specific pulses. Prey items that were removed from the diet following one day of feeding were only detected in scats collected within 48 h of ingestion. Proportions of fish DNA present in eight scat samples (evaluated through the screening of clone libraries) were roughly proportional to the mass of prey items consumed, raising the possibility that DNA quantification methods could provide semi-quantitative diet composition data. This study should be of broad interest to researchers studying diet since it highlights an approach that can accurately identify prey species and is not dependent on prey hard parts surviving digestion.

Genetic screening for prey in the gut contents from a giant squid (Architeuthis sp.).

Giant squids (Architeuthis sp.) remain mysterious; they have evaded observation and are rarely taken from their deep sea habitat. Information on the diet of Architeuthis is scarce due to the limited number of specimens with morphologically recognizable remains in their digestive tracts. We explored the use of polymerase chain reaction (PCR)-based methods for detection of DNA in the prey remains and amorphous slurry from an Architeuthis gut sample. The DNA region amplified varied in size, allowing separation of fish and squid components. Sequence comparisons identified fish prey as Macruronus novaezelandiae. Isolation of Architeuthis DNA from an ingested tentacle and the presence of chitin fragments indicate cannibalism occurs in giant squid. Denaturing gradient gel electrophoresis was used to screen for less common DNA types, revealing a high frequency of PCR-generated false alleles, but no additional prey species.

Group-specific polymerase chain reaction for DNA-based analysis of species diversity and identity in dietary samples.

Unique DNA sequences are present in all species and can be used as biomarkers for the detection of cells from that species. These DNA sequences can most easily be detected using the polymerase chain reaction (PCR), which allows very small quantities of target DNA sequence to be amplified even when the target is mixed with large amounts of nontarget DNA. PCR amplification of DNA markers that are present in a wide range of species has proven very useful for studies of species diversity in environmental samples. The taxonomic range of species to be identified from environmental samples may often need to be restricted to simplify downstream analyses and to ensure that less abundant sequences are amplified. Group-specific PCR primer sets are one means of specifying the range of taxa that produce an amplicon in a PCR. We have developed a range of group-specific PCR primers for studying the prey diversity found in predator stomach contents and scats. These primers, their design and their application to studying prey diversity and identity in predator diet are described.

A DNA-based method for identification of krill species and its application to analysing the diet of marine vertebrate predators.

Accurate identification of species that are consumed by vertebrate predators is necessary for understanding marine food webs. Morphological methods for identifying prey components after consumption often fail to make accurate identifications of invertebrates because prey morphology becomes damaged during capture, ingestion and digestion. Another disadvantage of morphological methods for prey identification is that they often involve sampling procedures that are disruptive for the predator, such as stomach flushing or lethal collection. We have developed a DNA-based method for identifying species of krill (Crustacea: Malacostraca), an enormously abundant group of invertebrates that are directly consumed by many groups of marine vertebrates. The DNA-based approach allows identification of krill species present in samples of vertebrate stomach contents, vomit, and, more importantly, faeces. Utilizing samples of faeces from vertebrate predators minimizes the impact of dietary studies on the subject animals. We demonstrate our method first on samples of Adelie penguin (Pygoscelis adeliae) stomach contents, where DNA-based species identification can be confirmed by prey morphology. We then apply the method to faeces of Adelie penguins and to faeces of the endangered pygmy blue whale (Balaenoptera musculus brevicauda). In each of these cases, krill species consumed by the predators could be identified from their DNA present in faeces or stomach contents.

Aerobic dive limit: how often does it occur in nature?

Diving animals offer a unique opportunity to study the importance of physiological constraint in their everyday behaviors. An important component of the physiological capability of any diving animal is its aerobic dive limit (ADL). The ADL has only been measured in a few species. The goal of this study was to estimate the aerobic dive limit from measurements of body oxygen stores and at sea metabolism. This calculated ADL (cADL) was then compared to measurements of diving behavior of individual animals of three species of otariids, the Antarctic fur seal, Arctocephalus gazella, the Australian sea lion, Neophoca cinerea, and the New Zealand sea lion, Phocarctos hookeri. Antarctic fur seals dove well within the cADL. In contrast, many individuals of both sea lion species exceeded the cADL, some by significant amounts. Australian sea lions typically dove 1.4 times longer than the cADL, while New Zealand sea lions on average dove 1.5 times longer than the cADL. The tendency to exceed the cADL was correlated with the dive pattern of individual animals. In both Antarctic Fur Seals and Australian sea lions, deeper diving females made longer dives that approached or exceeded the cADL (P<0.01, r(2)=0.54). Australian and New Zealand sea lions with longer bottom times also exceeded the cADL to a greater degree. The two sea lions forage on the benthos while the fur seals feed shallow in the water column. It appears that benthic foraging requires these animals to reach or exceed their aerobic dive limit.

Foraging energetics and diving behavior of lactating New Zealand sea lions, Phocarctos hookeri.

The New Zealand sea lion, Phocarctos hookeri, is the deepest- and longest-diving sea lion. We were interested in whether the diving ability of this animal was related to changes in its at-sea and diving metabolic rates. We measured the metabolic rate, water turnover and diving behavior of 12 lactating New Zealand sea lions at Sandy Bay, Enderby Island, Auckland Islands Group, New Zealand (50 degrees 30'S, 166 degrees 17'E), during January and February 1997 when their pups were between 1 and 2 months old. Metabolic rate (rate of CO(2) production) and water turnover were measured using the (18)O doubly-labeled water technique, and diving behavior was measured with time/depth recorders (TDRs). Mean total body water was 66.0+/-1.1 % (mean +/- s.d.) and mean rate of CO(2) production was 0. 835+/-0.114 ml g(-)(1 )h(-)(1), which provides an estimated mass-specific field metabolic rate (FMR) of 5.47+/-0.75 W kg(-)(1). After correction for time on shore, the at-sea FMR was estimated to be 6.65+/-1.09 W kg(-)(1), a value 5.8 times the predicted standard metabolic rate of a terrestrial animal of equal size. The mean maximum dive depth was 353+/-164 m, with a mean diving depth of 124+/-36 m. The mean maximum dive duration was 8.3+/-1.7 min, with an average duration of 3.4+/-0.6 min. The deepest, 550 m, and longest, 11.5 min, dives were made by the largest animal (155 kg). Our results indicate that the deep and long-duration diving ability of New Zealand sea lions is not due to a decreased diving metabolic rate. Individual sea lions that performed deeper dives had lower FMRs, which may result from the use of energetically efficient burst-and-glide locomotion. There are differences in the foraging patterns of deep and shallow divers that may reflect differences in surface swimming, time spent on the surface and/or diet. Our data indicate that, although New Zealand sea lions have increased their O(2) storage capacity, they do not, or cannot, significantly reduce their at-sea metabolic rates and are therefore likely to be operating near their physiological maximum.

Blood volume and diving ability of the New Zealand sea lion, Phocarctos hookeri.

We test the hypothesis that the New Zealand sea lion is physiologically better equipped for prolonged, continuous diving than other otariids (fur seals and sea lions) by measuring its blood volume, an important component of its oxygen storage. Mass, hematocrit, and plasma volume were measured and blood volume calculations were completed on 14 adult females and five juvenile females. Plasma volume was determined using the Evans blue dye dilution technique. Mean plasma volume for all subjects was 74 mL kg(-1). Mass-specific plasma volume was significantly higher in adult females (15.3%) than in juveniles (14.6%). Blood volume (150 mL kg(-1)) and hematocrit (51%) were not significantly different between adults and juveniles. The aerobic dive limit can be estimated by dividing the animal's oxygen stores by its metabolic rate. The estimated aerobic dive limit for adult animals was between 5.5 and 7.8 min, depending on the assumed metabolic rate. New Zealand sea lions have the highest blood volume yet reported for an otariid, which supports the hypothesis that they have a physiological capability suited to their unique diving behavior.

Evidence for a prolonged postimplantation period in the Australian sea lion (Neophoca cinerea).

Concentrations of circulating progesterone and oestradiol were measured in 96 free-ranging, female Australian sea lions Neophoca cinerea from Kangaroo Island, South Australia. There was a marked increase in the concentrations of both hormones (progesterone from approximately 12 ng ml-1 to approximately 24 ng ml-1; oestradiol from approximately 1.5 pg ml-1 to approximately 14 pg ml-1) about 3.5 months after the probable date of mating, reaching peak values in the 5 months after parturition. Progesterone concentrations remained at peak concentrations for about 2 months, decreasing at approximately 8 months to concentrations approximating those of the first 3 months after parturition. Oestradiol concentrations decreased, after reaching a peak, to 3-4 pg ml-1 at about 8 months after parturition. The timing of the increase in the concentrations of circulating progesterone and oestradiol provides evidence that the blastocyst reactivates and implants between 3.5 and 5 months of pregnancy in Australian sea lions, indicating an embryonic diapause of similar duration to that of other pinnipeds. This would suggest a prolonged postimplantation period of up to 14 months (to fit with the gestation period of 18 months reported for this species) the longest postimplantation period recorded for pregnancy in any pinniped.

Mass stranding of striped dolphin, Stenella coeruleoalba, at Augusta, Western Australia: notes on clinical pathology and general observations.

Seventeen striped dolphins, Stenella coeruleoalba, were found stranded on a West Australian beach. Three animals died before a rescue attempt was made and a further three died during the rescue. The remaining dolphins were released 24 km offshore and were not seen again. One dolphin was noted to have a broken mandible. Evidence of physical trauma to the other dolphins was minimal; one adult female was observed with some peeling skin. Blood was collected for analysis. All dolphins were slightly dehydrated and had a leukogram typical of a stressed animal. Plasma biochemistry reflected primary muscle trauma. There were no clues to the cause of the stranding; observed pathology reflected damage that occurred as a direct consequence of stranding.

Use of real-time B-mode ultrasound for pregnancy diagnosis and measurement of fetal growth rate in captive bottlenose dolphins (Tursiops truncatus).

Real-time ultrasonography was used to detect pregnancy in 4 captive bottle-nose dolphins. Pregnancy was readily confirmed from around the 4th month of gestation by imaging fetal fluids and fetal movement. Periodic examination permitted monitoring of the viability of the fetuses by observation of their heart beat and movement, and serial measurements of skull diameter (occipito-frontal axis) and thoracic diameter was possible. A growth curve for these measurements was plotted.

Prolonged and multiple immobilizations of the southern elephant seal using ketamine hydrochloride-xylazine hydrochloride or ketamine hydrochloride-diazepam combinations.

Thirty seven southern elephant seals (Mirounga leonina) were singularly or repeatedly immobilized with combinations of ketamine hydrochloride (HCl) and xylazine HCl or ketamine HCl and diazepam. Atropine sulphate was included in the drug combinations. To permit experimental procedures the seals were immobilized for periods of 30-330 min. The mean induction dose of ketamine HCl was 8.71 +/- 0.25 mg/kg (mean +/- SE). The mean induction time was 16.02 +/- 2.62 min. For the elephant seals immobilized for periods in excess of 180 min, the mean dose of ketamine HCl used per hr was 3.31 +/- 0.13 mg/kg/hr and the mean dose of ketamine HCl used per hr postinduction was 1.31 +/- 0.15 mg/kg/hr. The mean dose of diazepam used was 0.09 +/- 0.01 mg/kg and the mean dose of xylazine HCl was 0.41 +/- 0.01 mg/kg. Elephant seals were weighed on 20 occasions (weight range: 897-1,932 kg) and the relationship between standard length and weight was found to be: Weight = 9.98 length - 2,317.63 (r2 = 0.724). Adverse reactions to seals immobilized only once or twice were not observed. Two seals immobilized on three occasions developed abscesses at the site of injection.