Martina Đuras

Three Anisakis spp. isolated from toothed whales stranded along the eastern Adriatic Sea coast.

Knowledge concerning cetacean ecology in the Mediterranean is limited but important for sustainable planning and enforcement of appropriate conservation measures. Any information that might help to elucidate their ecology is essential. We explored the population and genetic structures of Anisakis spp. nematodes isolated from four toothed whale species - bottlenose dolphins (Tursiops truncatus), striped dolphins (Stenella coeruleoalba), Rissos dolphins (Grampus griseus) and Cuviers beaked whales (Ziphius cavirostris) - stranded along the eastern Adriatic Sea coast (1990-2012) to reveal more information on host ecological patterns. Lower parasite prevalence was observed in resident dolphin species compared with occasionally occurring species, as well as in young compared with adult dolphins, indicating different feeding habits related to age. No unequivocal relationship between the biological traits of a host (age, body length, body mass and blubber depth) and Anisakis population parameters was observed. Phylogenetic analysis revealed a new geographical record of Anisakis simplex sensu stricto (1.96%) and Anisakis physeteris (1.31%) in the Adriatic Sea in addition to resident Anisakis pegreffii (96.73%). In an assessment of the Adriatic Sea and oceans worldwide, the genetic structure of Anisakis revealed that A. pegreffii populations do not differ among various final host species but do differ with respect to geographical location in contrast to previously accepted Anisakis panmixia.

Evidence of Brucella strain ST27 in bottlenose dolphin (Tursiops truncatus) in Europe

Marine mammal brucellosis has been known for more than 20 years, but recent work suggests it is more widespread than originally thought. Brucella (B.) pinnipedialis has been isolated from pinnipeds, while B. ceti strains have been associated with cetaceans. Here we report a Brucella strain isolated from multiple lymph nodes of one bottlenose dolphin (Tursiops truncatus) during routine examination of dolphin carcasses found in the Croatian part of the northern Adriatic Sea during the summer of 2015. Classical bacteriological biotyping, PCR-based techniques (single, multiplex, PCR-RFLP) and 16S rRNA DNA sequencing were used to identify Brucella spp. Multiple-locus variable number tandem repeat analysis of 16 loci and multilocus sequence typing of 9 loci were used for genotyping and species determination. The combination of bacteriological, molecular and genotyping techniques identified our strain as ST27, previously identified as a human pathogen. This report provides, to our knowledge, the first evidence of ST27 in the Adriatic Sea in particular and in European waters in general. The zoonotic nature of the strain and its presence in the Adriatic, which is inhabited by bottlenose dolphins, suggest that the strain may pose a significant threat to human health.

HEMATOLOGICAL AND SERUM BIOCHEMICAL REFERENCE VALUES FOR THE POSAVINA AND CROATIAN COLDBLOOD HORSE BREEDS

Abstract In this study the hematological and biochemical parameters in the blood of indigenous Croatian working horse breeds were investigated. The Posavina and Croatian Coldblood horses are adapted to harsh environmental conditions and their blood parameters might differ from other horse breeds. The study was carried out on 100 mares and 12 stallions of ages from 2 to 19 years. Fifteen hematological and 19 biochemical parameters were analysed. Values of 22 parameters showed considerable overlapping with values obtained for other horse breeds, and substantial resemblance is evident with values reported for Pakistani working horses. Several reference values showed statistically significant effect of sex (Hb, ALB, CREA, TBIL and Ca where significantly higher in stallions; EOS, CK, ALP, UREA and Mg were significantly higher in mares). None of the parameters studied showed any differences associated with age. The adaptation of Posavina and Croatian Coldblood horses to the harsh environment of flooded pastures and the way of breeding might be reflected in their specific reference values. The obtained results enabled the establishment of hematological and biochemical reference values in the blood of Posavina and Croatian Coldblood horses that will in turn support clinical diagnosis and further research into horse physiology. Abstract U studiji je obavljeno ispitivanje hematoloških i biohemijskih parametara krvi kod radnih hrvatskih autohtonih rasa konja. Posavski i hrvatski hladnokrvnjak su dve rase konja koje su adaptirane na oštre uslove spoljašnje sredine pa se pretpostavlja da njihovi parametri krvi mogu da budu različiti u poređenju sa drugim rasama. Ispitivanje je obavljeno na 100 kobila i 12 pastuva starosti od 2 do 19 godina. Obavljena je analiza u odnosu na pedeset hematoloških i 19 biohemijskih parametara. Vrednosti koje su dobijene u odnosu na 22 parametra pokazale su značajna preklapanja u poređenju sa drugim rasama konja, pri čemu je ustanovljena značajna podudarnost sa vrednostima koje su karakteristične za pakistanskog radnog konja. Nekoliko referentnih vrednosti je pokazalo statistički značajnan utacaj pola (Hb, ALB, CREA, TBIL i Ca vrednosti su bile značajno veće kod pastuva; EOS, CK, ALP, UREA i Mg vrednosti su bile značajno veće kod kobila). Nijedan parametar koji je ispitivan nije pokazao bilo kakvu varijaciju u odnosu na starost životinja. Prilagođenost posavskog i hrvatskog hladnokrvnog konja na oštre i teške uslove spoljašnje sredine koji podrazumevaju ispašu na poplavljenim pašnjacima, kao i način uzgoja, može da ima posledice na specifične referentne vrednosti ispitivanih parametara. Dobijeni podaci omogućavaju definisanje hematoloških i biohemijskih referentnih vrednosti krvi posavske i hrvatske rase hladnokrvnih konja, a te vrednosti bi omogućavale kliničku dijagnostiku i buduća ispitivanja fiziologije konja.

Concentration of mercury and selenium concentrations in the tissues of five cetacean species from Croatian coastal waters

Mercury (Hg) and selenium (Se) concentrations were measured in muscle, liver, kidney, spleen and lung tissues of five cetacean species, three dolphin (Stenella coeruleoalba, Tursiops truncatus and Grampus griseus) and two whale species (Balaenoptera physalus and Ziphius cavirostris), stranded along the Croatian coast during the period 1999-2002. Statistically significant differences in Hg concentrations in muscle, spleen and lung, and Se in liver and lung of the different dolphin species were observed. Mercury levels in liver and spleen and Se levels in liver differed between young and adult T. truncatus species. A significant positive correlation between different tissue types for Hg and Se concentrations was observed. In all tissues tested, the lowest Hg and Se concentrations were found in B. physalus. Mercury concentrations were positively correlated with Se in all tissues. The results present one of few studies related to lung and spleen tissues in these mammals, particularly in the Adriatic Sea. Since very little data are available, this research provides new data on concentrations of Hg and Se in five cetacean species from the Adriatic Sea basin.

Variations in Pectoral Girdle Muscles in Dogs

Muscle variations take the form of additional muscle bundle or belly, unusual muscle origin or termination, as well as complete muscle absence. Knowledge of such variations not only has clinical importance for guiding surgery, but also can help reveal phylogenetic relationships. To improve our understanding of muscle variations in dogs, 57 medium‐sized, cross‐breed male and female adult animals were dissected as part of a gross anatomy course between 2005 and 2011. Variations in pectoral girdle muscles were observed in 7 (12.3%) dogs and took the form of an additional muscle bundle in the brachiocephalicus muscle (n = 2), in the omotransversarius muscle (n = 2), in the cervical part of the rhomboideus muscle (n = 2) and in the cervical part of the serratus ventralis muscle (n = 1). Muscle variation was bilateral in only one dog; it involved the omotransversarius muscle. The variations did not seem to be sex dependent. Such variations can appear regularly in dogs and should be taken into consideration during anatomical dissection.

Manganese Concentrations in Tissues and Skin of Three Dolphin Species Stranded in the Croatian Waters of the Adriatic Sea from 1995 to 2013

Manganese concentrations were determined in muscle, kidney, liver, spleen, lung and fat tissues and skin of three cetacean species, the bottlenose (Tursiops truncatus), striped (Stenella coeruleoalba) and Risso’s (Grampus griseus) dolphins, stranded along the Croatian Adriatic coast between 1995 and 2013. Mean ranges determined in tissues were (mg/kg, ww): muscle 0.23–0.27, liver 2.87–4.00, kidney 0.66–1.26, spleen 0.37–0.79, lung 0.18–0.57, skin 0.11–0.97, fat 0.06–0.42. The highest Mn mean levels in tissues were measured in species (mg/kg, ww): T. truncates lung 0.41, skin 0.97, fat 0.42; S. coeruleoalba muscle 0.26; G. griseus kidney 1.26, liver 4.00, spleen 0.64. Significant differences of Mn concentrations in the liver (p = 0.034), spleen (p = 0.037) and skin (p = 0.013) were found among the three dolphin species. Significant differences in Mn levels were found between young and adult T. truncates in kidney (p = 0.042), lung (p = 0.0040) and skin (p = 0.0034).

Whale Poaching Detection Based on Microscopic Characteristics of Bottlenose Dolphins’ (Tursiops truncatus) Bone Fragments

Although many countries prohibit whaling, it remains a significant cause of whale population decline. The meal of whales, likely killed during illegal whaling or caught accidentally, can ap-pear in a fish meal as a contaminant detectable by microscopic examination of bone fragments. In order to provide a rigorous basis for such a detection, microscopic characterization of bone fragments of 10 female and 10 male, less than a year to 21 years old bottlenose dolphins (Tur-siops truncatus), a widespread cetacean species herein used as a whale model, was performed and compared to the reference bone fragments of fish, ruminant, poultry and pig. The processing of bones mimicked that used in the production of meat and bone meals, while their description was based on qualitative characteristics, i.e. the shape of a bone fragment ; the shape, density and distribution of osteocyte lacunae, and the distribution and density of canaliculae. Bottlenose dol-phin bone fragments are smoothly contoured ; their elliptical osteocyte lacunae are clearly visible, while the canaliculae radiate from the lacunae in all directions. The comparison of these qualita-tive bone characteristics with that of other vertebrata under study revealed that bottlenose dol-phin bone fragments can definitely be differentiated from that of fish, may be differentiated from that of poultry and pig, but cannot be differentiated from that of ruminants. Measurements of the osteocyte lacunae showed the lacunar length to be strongly associated with animal’s age and lacunar shape, while their width was strongly associated with animal’s gender lacunar shape. The results indicate the possibility of detecting a whale meal admixture in a fish meal using light microscopy, which should be followed by PCR to enable the identification of the admixture source.

Bruceloza u morskih sisavaca, s posebnim osvrtom na Republiku Hrvatsku

After almost two decades of research and evidence of brucellosis in marine mammals, it has become widely known that this disease is prevalent in marine mammals on a global scale. Positive animals have been found in the northern Atlantic, Mediterranean Sea and in the Arctic, including the Barents Sea. Infected or exposed animals have been detected along the Atlantic and Pacific coasts of North America, off the coasts of Peru, Australia, New Zealand, Hawaii, the Solomon Islands, and in the Antarctic Ocean. Brucellosis most commonly occurs in the harbour porpoise (Phocoena phocoena), followed by the striped dolphin (Stenella coeruleoalba), Atlantic white-sided dolphin (Lagenorhynchus acutus), bottlenose dolphin (Tursiops truncatus), shortbeaked common dolphin (Delphinus delphis) and minke whale (Balaenoptera acutorostrata). A positive serological reaction has been confirmed in 35 species of whales and 14 species of seals, two subspecies of sea otters, one species of freshwater otter and the polar bear. To date, brucellosis has been found in marine mammals in the Mediterranean Sea in Italy and Spain. The first research on brucellosis in dolphins in the Republic of Croatia began in 2015. Brucella sp. was isolated from the lymph node of a bottlenose dolphin found dead in the Poreč region. This was the first evidence of brucellosis in dolphins in the Adriatic Sea. The isolated sample was identified as Brucella ceti strain ST27, making this the first record of this strain in Europe, which may present a significant threat to human health.

Whole-Genome Sequence of the First Sequence Type 27 Brucella ceti Strain Isolated from European Waters

ABSTRACT Brucella spp. that cause marine brucellosis are becoming more important, as the disease appears to be more widespread than originally thought. Here, we report a whole and annotated genome sequence of Brucella ceti CRO350, a sequence type 27 strain isolated from a bottlenose dolphin carcass found in the Croatian part of the northern Adriatic Sea.