Dominic J. Tollit

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.

Comparative distribution, movements and diet of harbour and grey seals from the Moray Firth, N.E. Scotland

The distribution, movements and foraging activity of harbour and grey seals from the inner Moray Firth, N.E. Scotland, were compared using a combination of observations at haul-out sites, VHF and satellite-link telemetry, and analyses of diet composition. Terrestrial abundance of both grey and harbour seals was highest during the summer but there were marked differences in the extent to which the two species moved between different haul-out sites. Harbour seals showed local seasonal changes in distribution, but all 21 radio-tagged seals moved only to alternative haul-out sites within a range of 75 km. In contrast, four of the five grey seals tracked from the Moray Firth moved to haul-out sites 125-365km away, and provided evidence of interchange between the Moray Firth and other grey seal breeding areas in Orkney, the Firth of Forth and the Farne Islands. All harbour seals foraged within 60 km of their haul-out sites, but showed seasonal variation in their foraging areas which was related to changes in their terrestrial distribution. In contrast, those grey seals which foraged within the Moray Firth travelled up to 145 km from haul-out sites. Nevertheless, there was some overlap in the foraging areas used by harbour seals and grey seals in more inshore areas. Sandeels, gadoids, flatfish and cephalopods formed over 95% of the diet of both species. However, dietary data, particularly for grey seals, may be biased toward those individuals which fed in more inshore areas. These results suggest that Moray Firth harbour seals can be considered as a relatively discrete population, with clear links between breeding, feeding and resting areas, and little exchange of adults between this and adjacent breeding areas in Orkney and the Tay Estuary. In contrast, grey seals from several different breeding sites appear to move into the Moray Firth in summer and use the area primarily for foraging and non-breeding haul-out.

Estimating harbour seal abundance and status in an estuarine habitat in north-east Scotland

1. Harbour seals are the most widespread of pinniped species, coming ashore onto a variety of different habitats to rest, moult and breed. Estimates of their abundance and status rely on counts of animals in terrestrial haul-out groups but it is not clear to what extent current techniques are appropriate for all habitats. 2. This study aimed to determine the most appropriate methods for estimating the abundance and status of harbour seals in an estuarine habitat in north-east Scotland. Regular low-tide counts were conducted to identify the best time for annual counts. Survey data for 1993 were then combined with telemetry data on seal activity to produce an estimate of abundance. Finally, simulations using data on the variability of counts within a single year were used to determine the power of these techniques to detect trends in abundance. 3. The results suggest that annual counts conducted during the pupping season (mid-June to mid-July) provide the best estimates of abundance in this habitat. These results contrast with those from studies in rocky-shore habitats where counts made during the August moult provided more reliable abundance estimates. 4. In 1993, an average of 1007 seals were hauled out at low tides during the period 15 June-15 July. There were significant differences in haul-out frequency for males (0.521 of low tides) and females (0.698 of low tides) but no within-sex variation was detected during this period. Combining the telemetry data with the results of counts from the Moray Firth produced an abundance estimate of 1653 (95% confidence limits 1471-1836). 5. The potential effects of within and between-year variation in haul-out behaviour are discussed in relation to identifying suitable indices of abundance for trend analysis. In view of likely seasonal changes in the sex structure of haul-out groups, we recommend that programmes to monitor trends in harbour seal abundance should conduct annual counts at two different points in the annual cycle.

Development and application of DNA techniques for validating and improving pinniped diet estimates

Polymerase chain reaction techniques were developed and applied to identify DNA from >40 species of prey contained in fecal (scat) soft-part matrix collected at terrestrial sites used by Steller sea lions (Eumetopias jubatus) in British Columbia and the eastern Aleutian Islands, Alaska. Sixty percent more fish and cephalopod prey were identified by morphological analyses of hard parts compared with DNA analysis of soft parts (hard parts identified higher relative proportions of Ammodytes sp., Cottidae, and certain Gadidae). DNA identified 213 prey occurrences, of which 75 (35%) were undetected by hard parts (mainly Salmonidae, Pleuronectidae, Elasmobranchii, and Cephalopoda), and thereby increased species occurrences by 22% overall and species richness in 44% of cases (when comparing 110 scats that amplified prey DNA). Prey composition was identical within only 20% of scats. Overall, diet composition derived from both identification techniques combined did not differ significantly from hard-part identification alone, suggesting that past scat-based diet studies have not missed major dietary components. However, significant differences in relative diet contributions across scats (as identified using the two techniques separately) reflect passage rate differences between hard and soft digesta material and highlight certain hypothesized limitations in conventional morphological-based methods (e.g., differences in resistance to digestion, hard part regurgitation, partial and secondary prey consumption), as well as potential technical issues (e.g., resolution of primer efficiency and sensitivity and scat subsampling protocols). DNA analysis of salmon occurrence (from scat soft-part matrix and 238 archived salmon hard parts) provided species-level taxonomic resolution that could not be obtained by morphological identification and showed that Steller sea lions were primarily consuming pink (Oncorhynchus gorbuscha) and chum (Oncorhynchus keta) salmon. Notably, DNA from Atlantic salmon (Salmo salar) that likely originated from a distant fish farm was also detected in two scats from one site in the eastern Aleutian Islands. Overall, molecular techniques are valuable for identifying prey in the fecal remains of marine predators. Combining DNA and hard-part identification will effectively alleviate certain predicted biases and will ultimately enhance measures of diet richness, fisheries interactions (especially salmon-related ones), and the ecological role of pinnipeds and other marine predators, to the benefit of marine wildlife conservationists and fisheries managers.

Proportion of prey consumed can be determined from faecal DNA using real‐time PCR

Reconstructing the diets of pinnipeds by visually identifying prey remains recovered in faecal samples is challenging because of differences in digestion and passage rates of hard parts. Analysing the soft‐matrix of faecal material using DNA‐based techniques is an alternative means to identify prey species consumed, but published techniques are largely nonquantitative, which limits their usefulness for some applications. We further developed and validated a real‐time PCR technique using species‐specific mitochondrial DNA primers to quantify the proportion of prey in the diets of Steller sea lions (Eumetopias jubatus), a pinniped species thought to be facing significant diet related challenges in the North Pacific. We first demonstrated that the proportions of prey tissue DNA in mixtures of DNA isolated from four prey species could be estimated within a margin of ∼12% of the percent in the mix. These prey species included herring Clupea palasii, eulachon Thaleichthyes pacificus, squid Loligo opalescens and rosethorn rockfish Sebastes helvomaculatus. We then applied real‐time PCR to DNA extracted from faecal samples obtained from Steller sea lions in captivity that were fed 11 different combinations of herring, eulachon, squid and Pacific ocean perch rockfish (Sebastes alutus), ranging from 7% to 75% contributions per meal (by wet weight). The difference between the average percentage estimated by real‐time PCR and the percentage of prey consumed was generally < 12% for all diets fed. Our findings indicate that real‐time PCR of faecal DNA can detect the approximate relative quantity of prey consumed for complex diets and prey species, including cephalopods and fish.

A comparison of techniques used to estimate body condition of southern elephant seals (Mirounga leonina)

The total body water (TBW) and body condition of 86 female southern elephant seals was estimated from tritiated water (HTO) dilution space analysis. HTO blood samples were analysed using two distillation methods (direct serum counts and evaporative freeze capture) that yielded significantly different estimates. Evaporative freeze capture is recommended for use because it is faster, cheaper, and provides a more precise TBW estimate of dilution space. Estimates of TBW were then compared with those derived from bioelectrical impedance analysis (BIA) and morphometric models. There were significant, positive relationships between TBW and BIA variables, but the level of accuracy was inadequate for BIA to be more useful than the other methods trialled. Morphometric models accurately estimated TBW (kg). Models developed from surface area (SA) (TBW = [SA * 82.58] – 86.94) and from a combination of mass (M), length (L), and girth (G) (TBW = [(M * 0.72) + (L * 5.49) + (G * 134.94) + 164.36)] provided the most accurate TBW estimates. In contrast, condition indices did not give accurate or reliable estimates of relative body condition.

Ship speed reduction is an effective mitigation for underwater noise effects

The Vancouver Fraser Port Authority’s ECHO Program led a voluntary 2-month ship slowdown trial during which 56% of piloted ships slowed down to <13 knots in a 16 nautical mile corridor. The goal was to determine if a slowdown could be used as a mitigation measure to reduce ship related underwater noise effects in core Southern Resident killer whale habitat. A calibrated hydrophone system was used to measure ambient noise levels from 10 Hz to 100 kHz during the trial and a representative 2-month baseline period. The hydrophone was located 2.3 (inbound) and 5 km (outbound) from the center of the shipping lanes at 23 m depth. Analyses of data with ships present showed a median broadband noise reduction of 2.5 dB. This reduction was highest in the 10–100 Hz decade band (3.1 dB) and lowest in the 10,000–100,000 Hz band (0.3 dB). A statistical model found that received noise levels were best described by the distance to ships, the presence of small boats, water velocity, slowdown period, ship speed through water, and wind speed. This highlights that appropriate temporal scales and the inclusion of covariate data are needed to adequately measure ship related changes in underwater ambient noise levels. The Vancouver Fraser Port Authority’s ECHO Program led a voluntary 2-month ship slowdown trial during which 56% of piloted ships slowed down to <13 knots in a 16 nautical mile corridor. The goal was to determine if a slowdown could be used as a mitigation measure to reduce ship related underwater noise effects in core Southern Resident killer whale habitat. A calibrated hydrophone system was used to measure ambient noise levels from 10 Hz to 100 kHz during the trial and a representative 2-month baseline period. The hydrophone was located 2.3 (inbound) and 5 km (outbound) from the center of the shipping lanes at 23 m depth. Analyses of data with ships present showed a median broadband noise reduction of 2.5 dB. This reduction was highest in the 10–100 Hz decade band (3.1 dB) and lowest in the 10,000–100,000 Hz band (0.3 dB). A statistical model found that received noise levels were best described by the distance to ships, the presence of small boats, water velocity, slowdown period, ship speed through wat...