Fatty acids in common minke whale (Balaenoptera acutorostrata) blubber reflect the feeding area and food selection,but also high endogenous metabolism

The fatty acid (FA) composition has been analysed in the blubber of 56 minke whales caught during the Norwegian commercial whaling period in 2009–2011. Minke whales from four regions were sampled: the North Sea, Vesterålen, Spitsbergen/Bear Island and Finnmark. The FA profiles of the whale blubber have been compared with FA profiles of potential prey species to investigate if FA analysis can be used to predict the diet of minke whales and how the FA profiles of the blubber reflect the regional ecosystem in which the whales were caught. Clear differences in blubber FA profiles were found between minke whales from different areas, and the results of the present study show that FA analysis of the blubber can be used to indicate the whale's diet, but there appears to be a strong impact from metabolism on several FAs. The whale blubber FAs are separate from those of the prey by having relatively high levels of FAs likely to originate from endogenous metabolism, such as 18:1n9 (Δ9-desaturation of 18:0); chain shortening products of 22:1(n-11); 20:1(n-11) and 18:1(n-11); as well as 22:5(n-3), which is an elongation product of 20:5(n-3). High metabolic activity in the adipose tissue was also evident by the clear stratification of FA profiles found throughout the blubber layer. It is remarkable that the whale blubber has much lower levels of the long-chain PUFAs 20:5(n-3) and 22:6(n-3) than found in the prey organisms. It is likely that this results from selective partitioning of diet FAs between the storage lipids and membrane lipids.     

How to tell a sea monster: molecular discrimination of large marine animals of the North Atlantic

Remains of large marine animals that wash onshore can be difficult to identify due to decomposition and loss of external body parts, and in consequence may be dubbed "sea monsters." DNA that survives in such carcasses can provide a basis of identification. One such creature washed ashore at St. Bernard's, Fortune Bay, Newfoundland, in August 2001. DNA was extracted from the carcass and enzymatically amplified by the polymerase chain reaction (PCR): the mitochondrial NADH2 DNA sequence was identified as that of a sperm whale (Physeter catodon). Amplification and sequencing of cryptozoological DNA with "universal" PCR primers with broad specificity to vertebrate taxa and comparison with species in the GenBank taxonomic database is an effective means of discriminating otherwise unidentifiable large marine creatures.     

BILIARY BILE ACID COMPOSITION OF THE PHYSETERIDAE (SPERM WHALES)

Abstract: The bile acid composition of bile obtained from the hepatopancreatic ducts of three species of sperm whales (Cetacea: Physeteridae) was investigated. Bile acids were isolated by adsorption chromatography and analyzed by sequential HPLC, SIMS, and GLC-MS. In each species the dominant bile acids were deoxycholic acid (a secondary bile acid formed by bacterial 7α-dehydroxylation of cholic acid), and chenodeoxycholic acid (a primary bile acid) which together composed more than 86% of biliary bile acids in all three species. In Physeter catodon (sperm whale) deoxycholic acid constituted 79%, and in Kogia breviceps (pygmy sperm whale) it was 61% of biliary bile acids. The sperm whale, which differs from other whales in having a remnant of a large intestine, is the second mammal identified to date in which deoxycholic acid is the predominant bile acid. The high proportion of deoxycholic acid indicates that in the Physeteridae, anaerobic fermentation occurs in its cecum, and that bile acids undergo enterohepatic cycling. Also found were minor proportions of cholic acid, as well as bacterial derivatives of chenodeoxycholic acid (ursodeoxycholic acid, lithocholic acid, and the 12β-epimer of allo-deoxycholic acid). Bile acids were conjugated with taurine in all species; however, in the sperm whale ( Physeter ) glycine conjugates were present in trace proportions. The bile acid hydroxylation pattern (12α- but not 6α-hydroxylation), lack of primary 5α- (allo) bile acids, and presence of glycine conjugated bile acids suggests the possibility that sperm whales originated from ancient artiodactyls.     

Nucleotide sequence of the D-loop region of the sperm whale (Physeter macrocephalus) mitochondrial genome

We have amplified, by the polymerase chain reaction, and have sequenced the D-loop region of the mitochondrial DNA from the sperm whale (Physeter macrocephalus). The sperm whale D-loop was aligned with D- loop sequences from four other cetaceans (Commerson's dolphin, orca, fin whale, and minke whale) and an out-group (cow). This alignment showed the sperm whale sequence to be larger than that of other cetaceans. In addition, some sequence blocks were highly conserved among all six species, suggesting roles in the functioning of mitochondrial DNA. Other blocks that were previously reported to be well conserved among cetaceans showed little sequence conservation with the sperm whale D-loop, which argues against the functional importance of these sequence blocks in cetaceans.      

Parasites in stranded cetaceans of Patagonia

There is an increasing interest in parasites of marine mammals of Argentina. Here, we examined several poorly known cetaceans, i.e., 2 spectacled porpoises and 1 Burmeister's porpoise (Phocoenidae), and 1 Gray's beaked whale and 1 Cuvier's beaked whale (Ziphidae); we also updated the parasite information for 1 sperm whale (Physeteridae). These hosts strand only occasionally. We found Anisakis simplex s.l. in 2 spectacled porpoises and the Burmeister's porpoise, and recorded its distribution among the stomach chambers. Anisakis physeteris infected the sperm whale; Corynosoma cetaceum occurred in the duodenal ampulla of the Burmeister's porpoise; Corynosoma australe was found in the posterior-most region of the intestine of 1 spectacled porpoise, while another one had Tetrabothrius sp. in the anal crypts; Corynosoma bullosum and Corynosoma sp. were found in the sperm whale. The only digenean found was Pholeter gastrophilus in the Burmeister's porpoise. Merocercoids of Phyllobothrium delphini were present in the blubber of 1 spectacled porpoise, the sperm whale, and the Gray's beaked whale, while Scolex pleuronectis infected the Gray's beaked whale and 1 spectacled porpoise. No parasites were recovered from the Cuvier's beaked whale. Poor parasite-species assemblages are consistent in marine mammals of Patagonia. Given the conservation status of these hosts, the limited parasitological information gathered is valuable for conservation or management of these hosts in Patagonia.     

High concentrations of isovaleric acid in the fats of odontocetes: variation and patterns of accumulation in blubber vs. stability in the melon

Isovaleric acid (iso5:0) is an unusual fatty acid that is important for echolocation and hearing in acoustic tissues of some odontocetes, but its functional significance in blubber is unknown. We examined patterns of accumulation of this compound in blubber in 30 species of odontocetes ( n=299). Iso5:0 concentrations in blubber varied with phylogeny, ontogeny and body topography. Iso5:0 accumulated in greater quantities in superficial/outer blubber than in deep/inner blubber. In the outer blubber of northern right whale and Hector's dolphins, iso5:0 accounted for one-third to one-half of all fatty acids. Total blubber burden of iso5:0 in harbour porpoises represented up to 15 times the amount deposited in the melon. The composition of the melon does not change during starvation in harbour porpoises, supporting the hypothesis that lipids in melon are conserved for a specific function. Some odontocetes continually deposit iso5:0 in blubber after levels in melon have reached asymptotic levels, suggesting independent control of iso5:0 synthesis and storage in these compartments. Dolphins and porpoises inhabiting cold waters possess higher concentrations of iso5:0 in their outer blubber layers than species from warmer regions. We propose that this relationship represents an adaptive secondary role for iso5:0 in maintaining blubber flexibility in cold environments.     

Molecular studies on two variant repeat types of the common cetacean DNA satellite of the sperm whale, and the relationship between Physeteridae (sperm whales) and Ziphiidae (beaked whales)

In the sperm whale (Physeter macrocephalus) two different repeat types (A and B) of the common cetacean DNA satellite were identified. The evolution of each group of repeats appears to be independent from that of the other. The sequence similarity between the two groups is less than the similarity between group A and repeats of the satellite in related whale species. The systematic relationship within and between the families Physeteridae (sperm whales) and Ziphiidae (beaked whales) was addressed by both sequence analysis of the satellite and comparisons with the families Delphinidae and Phocoenidae. The mysticete blue whale (Balaenoptera musculus) was used as an outgroup in the comparisons. The molecular phylogeny, when maximum-parsimony analysis and the neighbor-joining method were used, grouped together species of each family. At the family level the ziphiids grouped closet to the families Phocoenidae and Delphinidae. The similarities between the common cetacean satellite of the blue whale and the sperm whale were greater than those between the blue whale and the other odontocetes included, suggesting that the evolution of the satellite is slower in the sperm whale than in the other odontocetes.      

WEIGHTS AND ANATOMICAL MEASUREMENTS OF NORTHEASTERN ATLANTIC FIN (BALAENOPTERA PHYSALUS, LINNAEUS) AND SEI (B. BOREALIS, LESSON) WHALES

Piecemeal body weights of eleven fin and four sei whales and intact weights of three foetuses, obtained from Iceland, are compared with published weight data. The Icelandic fin are similar to other northern hemisphere animals but are significantly leaner than their Antarctic counterparts. The Icelandic sei appear heavier than the North Pacific sei whales. Their weights cannot be predicted from a North Pacific sei whale weight/length formula. Length, girth and blubber thickness measurements indicate changes in relative body dimensions in the early fin whale foetus compared with juveniles and adults; however, the midterm sei whale foetus is similar to the adult and juvenile sei whales. The blubber appears to form a major component even in the foetal body. The integration of a standard series of lengths, girths and blubber thicknesses in juveniles and adults can provide an estimate of the blubber component. Both girth and length are significant parameters in estimating body weight, a simple weight/length formula being found to be inadequate to allow for variability in body fatness. Evaluation of such a multiple parameter formula for calculating weight appears satisfactory for both fin and sei whales. Apparent weight/length differences between species and stocks may thus be partly due to variations in body fatness.     

A primitive myoglobin from Tetrahymena pyriformis: its heme environment, autoxidizability, and genomic DNA structure

A myoglobin-like protein isolated from Tetrahymena pyriformis is composed of 121 amino acid residues. This is much smaller than sperm whale myoglobin by 32 residues, suggesting a distinct origin from the common globin gene. We have therefore examined this unique protein for its structural, spectral and stability properties. As a result, the rate of autoxidation of Tetrahymena oxymyoglobin (MbO(2)) was found to be almost comparable to that of sperm whale MbO(2) over a wide range of pH 4-12 in 0.1 M buffer at 25 degrees C. Moreover, both pH profiles exhibited the remarkable proton-assisted process, which can be performed in sperm whale myoglobin by the distal (E7) histidine as its catalytic residue. These kinetic observations are also in full accord with spectral examinations for the presence of a distal histidine in ciliated protozoa myoglobin. At the same time, we have isolated the globin genes both from T. pyriformis and Tetrahymena thermophila, and found that there is no intron in their genomic structures. This is in sharp contrast to previous reports on the homologous globin genes from Paramecium caudatum and Chlamydomonas eugametos. Rather, the Tetrahymena genes seemed to be related to the cyanobacterial globin gene from Nostoc commune. These contracted or truncated globins thus have a marked diversity in the cDNA, protein, and genomic structures.     

Patterns of blubber fat deposition and evaluation of body condition in growing southern right whale calves (Eubalaena australis)

Marine mammals rely on blubber mainly for energy storage, buoyancy, and streamlining. Mysticetes are born with a relatively thin fat layer that grows rapidly during nursing. However, little information on blubber deposition patterns is available for baleen whale calves. We measured blubber thickness at nine body locations in 350 southern right whale (Eubalaena australis) newborn to 4–6-month-old calves that died on the Península Valdés (Argentina) calving ground from 2003 to 2019, to document changes in blubber thickness with growth. Additionally, we looked for differences in blubber thickness and lipid content of the outer/superficial blubber in calves that died in years with high (2003, 2005, 2007–2013) and low calf mortality (2004, 2006, 2014–2019) to test whether the former were suffering from gross nutritional stress. Blubber thickness increased at all body locations with calf length. Along the cranio-caudal axis, blubber increased in the dorsal and ventral planes, but decreased laterally towards the peduncle, possibly to improve streamlining. We found no difference in blubber thickness and lipid content between high and low mortality years, suggesting that individuals were not undernourished. This is the first study to describe progressive increases in calf blubber during growth and contributes knowledge to right whale health and ontogeny.     

Use of PCR and partial sequencing of the large-subunit rRNA gene to identify Alexandrium catenella (Dinophyceae) from the South of Chile

A fragment of the large-subunit ribosomal DNA gene (LSU rDNA) from Chilean Alexandrium catenella clones isolated from two different geographic regions (XI and XII) was amplified by PCR and the products cloned and sequenced. Based on the analysis of the PCR products it is possible to distinguish two strains of A. catenella, denominated strain type 1 (a single PCR product band) and strain type 2 (two PCR product bands). These two strains proliferate in both, the XI and XII regions. Only in the XI region, there is evidence that they bloom simultaneously. The LSU rDNA sequence analysis indicate that the Chilean A. catenella isolated clones are more related to the North American ribotype-Western subribotype.     

Rope trauma,sedation, disentanglement,and monitoring‐tag associated lesions in a terminally entangled North Atlantic right whale (Eubalaena glacialis)

A chronically entangled North Atlantic right whale, with consequent emaciation was sedated, disentangled to the extent possible, administered antibiotics, and satellite tag tracked for six subsequent days. It was found dead 11 d after the tag ceased transmission. Chronic constrictive deep rope lacerations and emaciation were found to be the proximate cause of death, which may have ultimately involved shark predation. A broadhead cutter and a spring‐loaded knife used for disentanglement were found to induce moderate wounds to the skin and blubber. The telemetry tag, with two barbed shafts partially penetrating the blubber was shed, leaving barbs embedded with localized histological reaction. One of four darts administered shed the barrel, but the needle was found postmortem in the whale with an 80º bend at the blubber‐muscle interface. This bend occurred due to epaxial muscle movement relative to the overlying blubber, with resultant necrosis and cavitation of underlying muscle. This suggests that rigid, implanted devices that span the cetacean blubber muscle interface, where the muscle moves relative to the blubber, could have secondary health impacts. Thus we encourage efforts to develop new tag telemetry systems that do not penetrate the subdermal sheath, but still remain attached for many months.     

The complete amino acid sequence of the major component myoglobin of Amazon river dolphin (Inia geoffrensis).

The complete amino acid sequence of the major component myoglobin from Amazon River dolphin, Inia geoffrensis, was determined by specific cleavage of the protein to obtain large peptides which are readily degraded by the automatic sequencer. Three easily separable peptides were obtained by cleaving the protein with cyanogen bromide at the methionine residues and four peptides were obtained by cleaving the methyl-acetimidated protein with trypsin at the arginine residues. From these peptides over 85% of the sequence was completed. The remainder of the sequence was obtained by fragmentation of the large cyanogen bromide peptide with trypsin. This protein differs from that of the common porpoise, Phocoena phocoena, at seven positions, from that of the common dolphin, Delphinus delphis, at 11 positions, and from that of the sperm whale, Physeter catodon, at 15 positions. By comparison of this sequence with the three-dimensional structure of sperm whale myoglobin it appears that those residues close to the heme group are most conserved followed by those in nonhelical regions and lastly by those in the helical segments. All of the substitutions observed in this sequence fit easily into the three-dimensional structure of the sperm whale myoglobin.     

Cytoplasmic and Mitochondrial Creatine Kinases from the Skeletal Muscle of Sperm Whale (Physeter macrocephalus). Molecular Cloning and Enzyme Characterization

We have amplified two cDNAs, coding for creatine kinases (CKs), from the skeletal muscle of sperm whale Physeter macrocephalus by PCR, and cloned these cDNAs into pMAL plasmid. These are the first CK cDNA and deduced amino acid sequences from cetaceans to be reported. One of the two amino acid sequences is a cytoplasmic, muscle-type isoform (MCK), while the other was identified as a sarcomeric, mitochondrial isoform (sMiCK) that included a mitochondrial targeting peptide. The amino acid sequences of sperm whale MCK and sMiCK showed 94–96% sequence identity with corresponding isoforms of mammalian CKs, and all of the key residues necessary for CK function were conserved. The phylogenetic analyses of vertebrate CKs with three independent methods (neighbor-joining, maximum-likelihood and Bayes) supported the clustering of sperm whale MCK with Bos and Sus MCKs, in agreement with the contemporary view that these groups are closely related. Sperm whale MCK and sMiCK were expressed in Escherichia coli as a fusion protein with maltose-binding protein, and the kinetic constants (K _m, K _d and k _cat) were determined for the forward reaction. Comparison of kinetic constants with those of human and mouse CKs indicated that sperm whale MCK has a comparable affinity for creatine (K _m^Cr = 9.38 mM) to that of human MCK, and the sMiCK has two times higher affinity for creatine than the human enzyme. Both the MCK and sMiCK of sperm whale display a synergistic substrate binding (K _d /K _m = 3.1–7.8) like those of other mammalian CKs.     

Characterization of Fasciola spp. in Myanmar on the basis of spermatogenesis status and nuclear and mitochondrial DNA markers

Fasciola spp. in Myanmar were characterized on the basis of spermatogenesis status and DNA markers of nuclear internal transcribed spacer 1 (ITS1) and mitochondrial NADH dehydrogenase subunit 1 (nad1). We collected 88 adult flukes from Yangon, Lashio, and Myitkyina. Spermatogenesis status was analyzed by the presence of sperm in the seminal vesicles, and 8 aspermic and 80 spermic flukes were detected. The flukes were identified on the basis of spermatogenesis status and ITS1 types which were analyzed by a PCR-RFLP method, and 80 spermic flukes were identified as F. gigantica. A very low detection rate of aspermic Fasciola sp. indicated that they are not established in Myanmar. In phylogenetic analyses, the 7 aspermic Fasciola sp. from Myitkyina displayed a haplotype in nad1 sequence, which was identical to that of aspermic Fasciola sp. from other Asian countries including China. Therefore, they were probably introduced from China through an infected domestic ruminant. On the other hand, 17 nad1 haplotypes detected in F. gigantica belonged to 2 clades unique to Myanmar, each with a distinct founder haplotype in a network analysis. This indicated a unique history of F. gigantica introduction into Myanmar involving ancient artificial movements of domestic ruminants.     

A rapid method for positioning small flexible molecules, nucleic acids, and large protein fragments in experimental electron density maps.

A Monte Carlo procedure, encoded in the program Blob, has been developed and tested for the purpose of positioning large molecular fragments or small flexible molecules in electron density maps. The search performed by the algorithm appears to be sufficiently thorough to accurately position a small flexible ligand in well-defined density while remaining sufficiently random to offer interesting alternate suggestions for density representing disordered binding modes of a ligand. Furthermore, the algorithm is shown to be efficient enough to accurately position large rigid molecular fragments. In the first of the test cases with large molecular fragments, Blob was surprisingly effective in positioning a poly-alanine model of a 53-residue domain in poor electron density resulting from molecular replacement with a partial model. At 3.0 A resolution the domain was positioned consistently within 0.2 A of its experimentally determined position. Even at 6.0 A resolution Blob could consistently position the domain to within 0.75 A of its actual position. A second set of tests with large molecular fragments revealed that Blob could correctly position large molecular fragments with quite significant deviations from the actual structure. In this test case, fragments ranging from a 170-residue protein domain with a 3.8 A rms deviation from the actual structure to a 22-base pair ideal B-form DNA duplex were positioned accurately in a 3.2 A electron density map derived from multiple isomorphous replacement methods. Even when decreasing the quality of the maps, from a figure of merit of 0.57 to as low as 0. 35, Blob could still effectively position the large protein domain and the DNA duplex. Since it is efficient, can handle large molecular fragments, and works in poor and low resolution maps, Blob could be a useful tool for interpreting electron density maps in de novo structure determinations and in molecular replacement studies. Proteins 1999;36:512-525.     

Complete amino acid sequence of the major component myoglobin of finback whale (Balaenoptera physalus)

The complete amino acid sequence of the major component myoglobin from finback whale, Balaenoptera physalus, was determined by the automated Edman degradation of several large peptides obtained by specific cleavages of the protein. Three easily separable peptides were obtained by cleaving with cyanogen bromide at the two methionine residues and one large peptide was isolated after cleavage with (2-p-nitrophenylsulfenyl)-3-methyl-3'-bromoindolenine. More than 60% of the covalent structure was established by the sequential degradation of three of these peptides and the apomyoglobin. An additional 30% of the primary sequence was established with peptides obtained from tryptic digestion of both the apomyoglobin and the acetimidoapomyoglobin, and the final 10% of the sequence was completed after digestion of the two larger cyanogen bromide peptides with S. aureus strain V8 protease. This myoglobin differs from that of the sperm whale, Physeter catodon, at 15 positions, from that of the arctic minke whale, Balaenoptera acutorostrata, at 3 positions, and from that of the California gray whale, Eschrichtius gibbosus, at 4 positions. All of the substitutions observed in this sequence fit easily into the three-dimensional structure of the sperm whale myoglobin.     

Blubber fatty acid composition of bowhead whales, Balaena mysticetus: Implications for diet assessment and ecosystem monitoring

Fatty acids (FA) have a diversity of structures that are transferred with little modification through food webs, making them valuable in assessing diets of animals that cannot be directly observed feeding. Before using FA to estimate diets, it is necessary to evaluate variation in FA signatures within and among individuals of a given species. To begin assessing diets and foraging of western Arctic bowhead whales (Balaena mysticetus), we examined the FA in blubber of 64 bowheads taken in the spring and fall subsistence hunts in 1997–2002 at Barrow and Kaktovik, Alaska. We found no significant differences in FA characteristics of inner blubber layers taken from either duplicate samples on the dorsal surface, or between dorsal and ventral sites. Significant differences were found in the FA composition between inner and outer layers of blubber at the same body site. We also found age, season and year to have significant effects on FA composition; however, gender was not found to be significant. While the importance of the Beaufort Sea as a feeding ground of bowhead whales remains uncertain, our results indicate that adults and sub-adults foraged to some extent on different prey and that both age classes consumed copepods there in summer at sufficient levels to significantly alter their blubber FA profiles. Both of these findings correspond with dietary conclusions reached from the analysis of stomach contents. Furthermore, we found compelling evidence that yearly variation in bowhead FA reflect changes in FA compositions of phytoplankton at the base of the food web, probably in response to climate variation. Variability in phytoplankton-derived FA in blubber was correlated significantly with yearly mean values of the Pacific Decadal Oscillation. FA in bowhead whale blubber, therefore, might be used to monitor effects of climate change on lower trophic levels and production processes in the western Arctic.     

Molecular identification of novel alpha- and gammaherpesviruses from cetaceans stranded on Japanese coasts

Herpesviral infections have been documented in some cetaceans; however, they have not yet been identified in species in the western North Pacific. In the present study, 178 tissue samples from 76 stranded cetacean individuals were tested for the presence of herpesviruses. Herpesvirus genomic DNA fragments surrounding the DNA polymerase gene were amplified in samples from four individuals. TA cloning and direct sequencing of these DNA fragments revealed the presence of two novel alphaherpesviruses, and two novel gammaherpesviruses in the four cetacean individuals. The alphaherpesviruses were associated with the lung tissue of a false killer whale (Pseudorca crassidens), and with the mucus of a melon-headed whale (Peponocephala electra). The gammaherpesviruses were found in the lymph tissues of a Stejneger's beaked whale (Mesoplodon stejnegeri) and a sperm whale (Physeter macrocephalus). The phylogenetic tree using amino acid sequences of the DNA polymerase gene supported the inclusion of the novel viruses identified here in a single monophyletic group containing alphaherpesviruses from other Atlantic cetacean species. Conversely, the novel gammaherpesviruses formed an independent clade distant from other known cetacean gammaherpesviruses.