Transient expression of rabies virus G-glycoprotein using BHK-21 cells cultured in suspension

Objective To assess the expression of rabies virus G-glycoprotein (RVGP) expression using Semliki Forest virus as a vector in combination with BHK-21 cells cultured in suspension. Results A multilevel factorial design was used to quantify effects of temperature (33–37 C), fresh medium addition after the viral adsorption step (100–200 % with respect to the initial cell suspension volume before infection) and harvest time (8–40 h) on RVGP production. Experimental runs were performed in 24-well cell culture plates at a multiplicity of infection (MOI) of 16. An additional experiment in spinner-flask was performed at MOI of 9, using the optimal conditions determined in cell culture plates. Values for temperature, fresh medium addition and harvest time of 33 C, 100 % and 16 h, respectively, ensured the optimal RVGP production in culture plates. The volumetric yield (239 ng ml-1 ) in these conditions was higher than that reported previously for adherent cell culture. In spinner-flasks, the volumetric yield was improved (559 ng ml-1 ). Conclusion These results establish the basis for designing bioprocess to produce RVGP.

Cone-beam evaluation of pharyngeal airway space in class I, II, and III patients

The purpose of this study is to make a 3-dimensional (3-D) evaluation of the pharyngeal airway space (PAS) in patients with class I, II, and III malocclusion. Sixty patients were evaluated. The patients were divided in 3 groups according to their occlusion classification. The volume and area of PAS were evaluated using the software Dolphin 3-D Imaging in the preoperative period for orthognathic surgery. PAS volume and area were influenced by different patterns of malocclusion. The mean volume and area for class III patients were statistically bigger than for classes I and II patients (P < .001). There was also a significant difference for volume values between class I and II patients, being the bigger volume for the class I patients (P < .05). It was possible to conclude that the class III patients presented a bigger PAS compared with classes I and II patients.

Toxic heritage: maternal transfer of pyrethroid insecticides and sunscreen agents in dolphins from Brazil

Pyrethroids (PYR) and UV filters (UVF) were investigated in tissues of paired mother-fetus dolphins from Brazilian coast in order to investigate the possibility of maternal transfer of these emerging contaminants. Comparison of PYR and UVF concentrations in maternal and fetal blubber revealed Franciscana transferred efficiently both contaminants to fetuses (F/M > 1) and Guiana dolphin transferred efficiently PYR to fetuses (F/M > 1) different than UVF (F/M < 1). PYR and UVF concentrations in fetuses were the highest-ever reported in biota (up to 6640 and 11,530 ng/g lw, respectively). Muscle was the organ with the highest PYR and UVF concentrations (p < 0.001), suggesting that these two classes of emerging contaminants may have more affinity for proteins than for lipids. The high PYR and UVF concentrations found in fetuses demonstrate these compounds are efficiently transferred through placenta. This study is the first to report maternal transfer of pyrethroids and UV filters in marine mammals.

Genetic isolation and morphological divergence of Black Sea bottlenose dolphins

The Black Sea is a semi-enclosed body of water that differs from the adjacent Mediterranean Sea in terms of its biodiversity, oceanographical and ecological characteristics. There is growing international concern about pollution in the Black Sea and other anthropogenic threats to its fauna. The bottlenose dolphin (Tursiops truncatus) is one of three species of cetaceans living in the Azov-Black Sea basin. Despite considerable research on bottlenose dolphins elsewhere, the extent of human impacts on the Black Sea populations is unknown. Previous attempts to award special conservation status to Black Sea cetaceans have failed specifically because policy makers have viewed their ecological and evolutionary uniqueness as equivocal. This study assessed divergence between Black Sea, Mediterranean Sea and Atlantic Ocean bottlenose dolphins for 26 cranial measurements (n = 75 adult bottlenose dolphin skulls) and mitochondrial DNA (n = 99 individuals). Black Sea bottlenose dolphins are smaller than those in the Mediterranean, and possess a uniquely shaped skull. As in a previous study, we found the Black Sea population to be genetically distinct, with relatively low levels of mtDNA diversity. Population genetic models suggest that Black Sea bottlenose dolphins have so little gene flow with the Mediterranean due to historical isolation that they should be managed separately.

Saving the Vaquita: Immediate Action, Not More Data

The recent likely extinction of the baiji (Chinese river dolphin [Lipotes vexillifer]) (Turvey et al. 2007) makes the vaquita (Gulf of California porpoise [Phocoena sinus]) the most endangered cetacean. The vaquita has the smallest range of any porpoise, dolphin, or whale and, like the baiji, has long been threatened primarily by accidental deaths in fishing gear (bycatch) (Rojas-Bracho et al. 2006). Despite repeated recommendations from scientific bodies and conservation organizations, no effective actions have been taken to remove nets from the vaquita’s environment. Here, we address three questions that are important to vaquita conservation: (1) How many vaquitas remain? (2) How much time is left to find a solution to the bycatch problem? and (3) Are further abundance surveys or bycatch estimates needed to justify the immediate removal of all entangling nets from the range of the vaquita? Our answers are, in short: (1) there are about 150 vaquitas left, (2) there are at most 2 years within which to find a solution, and (3) further abundance surveys or bycatch estimates are not needed. The answers to the first two questions make clear that action is needed now, whereas the answer to the last question removes the excuse of uncertainty as a delay tactic. Herein we explain our reasoning.

Taxonomy of the Common Dolphins of the Eastern Pacific Ocean

Delphinus bairdii Dall is a species of dolphin distinct from D. delphis Linnaeus, with which it has usually been synonymized. D. bairdii has a longer rostrum relative to the zygomatic width of the skull; the ratio of these measurements falls at 1.55 or above for bairdii and 1.53 and below for delphis. In the eastern Pacific Ocean, D. bairdii is found in the Gulf of California and along the west coast of Baja California, Mexico; D. delphis is presently found in the waters off California. Until approximately the beginning of the present century, bairdii occurred farther north in the eastern Pacific Ocean, at least to the Monterey Bay area of California. Restriction of bairdii to more southerly waters, probably as an indirect result of a change in water temperature, may have permitted delphis to move into inshore Californian waters. The Pacific population of D. delphis has a somewhat shorter rostrum than the Atlantic population, and is perhaps subspecifically different. A thorough analysis of the entire genus Delphinus is needed before the relationship of all the populations can be understood and names properly applied.

Past and present distribution, densities and movements of blue whales Balaenoptera musculus in the Southern Hemisphere and northern Indian Ocean

1. Blue whale locations in the Southern Hemisphere and northern Indian Ocean were obtained from catches (303 239), sightings (4383 records of ≥ 8058 whales), strandings (103), Discovery marks (2191) and recoveries (95), and acoustic recordings. 2. Sighting surveys included 7 480 450 km of effort plus 14 676 days with unmeasured effort. Groups usually consisted of solitary whales (65.2%) or pairs (24.6%); larger feeding aggregations of unassociated individuals were only rarely observed. Sighting rates (groups per 1000 km from many platform types) varied by four orders of magnitude and were lowest in the waters of Brazil, South Africa, the eastern tropical Pacific, Antarctica and South Georgia; higher in the Subantarctic and Peru; and highest around Indonesia, Sri Lanka, Chile, southern Australia and south of Madagascar. 3. Blue whales avoid the oligotrophic central gyres of the Indian, Pacific and Atlantic Oceans, but are more common where phytoplankton densities are high, and where there are dynamic oceanographic processes like upwelling and frontal meandering. 4. Compared with historical catches, the Antarctic (‘true’) subspecies is exceedingly rare and usually concentrated closer to the summer pack ice. In summer they are found throughout the Antarctic; in winter they migrate to southern Africa (although recent sightings there are rare) and to other northerly locations (based on acoustics), although some overwinter in the Antarctic. 5. Pygmy blue whales are found around the Indian Ocean and from southern Australia to New Zealand. At least four groupings are evident: northern Indian Ocean, from Madagascar to the Subantarctic, Indonesia to western and southern Australia, and from New Zealand northwards to the equator. Sighting rates are typically much higher than for Antarctic blue whales.

Evolution of river dolphins

The world's river dolphins (Inia, Pontoporia, Lipotes and Platanista) are among the least known and most endangered of all cetaceans. The four extant genera inhabit geographically disjunct river systems and exhibit highly modified morphologies, leading many cetologists to regard river dolphins as an unnatural group. Numerous arrangements have been proposed for their phylogenetic relationships to one another and to other odontocete cetaceans. These alternative views strongly affect the biogeographical and evolu- tionary implications raised by the important, although limited, fossil record of river dolphins. We present a hypothesis of river dolphin relationships based on phylogenetic analysis of three mitochondrial genes for 29 cetacean species, concluding that the four genera represent three separate, ancient branches in odontocete evolution. Our molecular phylogeny corresponds well with the first fossil appearances of the primary lineages of modern odontocetes. Integrating relevant events in Tertiary palaeoceanography, we develop a scenario for river dolphin evolution during the globally high sea levels of the Middle Miocene. We suggest that ancestors of the four extant river dolphin lineages colonized the shallow epicontinental seas that inundated the Amazon, Parana, Yangtze and Indo-Gangetic river basins, subsequently remaining in these extensive waterways during their transition to freshwater with the Late Neogene trend of sea-level lowering.

Proposed Updates to the List of Recognised Species of Cetaceans

Addition of three species to the list is recommended based on recent literature. (Orcaella brevirostris) has been split into the Irrawaddy dolphin (O. brevirostris) and the Australian snubfin dolphin (O. heinsohni). Sotalia fluviatilis has been split into the riverine tucuxi (S. fluviatilis) and the marine "costero" (S. guianensis). Evidence to support both of these splits is convincing, and we recommend that they be recognized in the list. The existence of the Bryde's-whale-like species described in 2003 as Balaenoptera omurai has been confirmed with additional genetic (nuclear) data. While the species clearly exists, the nomenclature is still unsettled because the genetic identity of the holotype specimen of Balaenoptera edeni has not yet been determined. However, the name B. omurai is gaining wide usage in application to the new species, and we propose that it be used provisionally by the Scientific Committee pending the genetic identification of the B. edeni holotype. We recommend that India be urged to facilitate the identification. We recommend continued use of the name Balaenoptera edeni provisionally for both the "ordinary" large form and the small coastal form, recognizing that further genetic and morphological research may justify recognition of two species: B. brydei and B. edeni. We also recommend that any new specimen be referred to B. omurai only after its mtDNA has been sequenced and found to support the identification.

Levels of persistent organic pollutants and residual pattern of DDTs in small cetaceans from the coast of Sao Paulo, Brazil

The State of Sao Paulo is the most developed area in Brazil and was impacted by persistent organic pollutants for several decades. This study investigated organochlorines in five species of small cetaceans (Pontoporia blainvillei, Stenella frontalis, Sotalia guianensis, Tursiops truncatus and Steno bredanensis) found dead along the coast of Sao Paulo between 1997 and 2003. DDTs (15.9 mu g g(-1) lipid: mean for all pooled individuals) and PCBs (8.08 mu g g(-1)) exhibited the highest concentrations in the animals, reflecting large amounts formerly used in Brazil. Lower levels of mirex (0.149 mu g g(-1)), HCB (0.051 mu g g(-1)), CHLs (0.008 mu g g(-1)) and HCHs (0.007 mu g g(-1)) were detected in all species. Residual pattern of DDTs in dolphins suggests that o,p`-DDT is more recalcitrant than p,p`-DDT in the body of the animals and/or the environment. In contrast to p,p`-DDT, residues of o,p`-DDT seem to be preferentially converted into o,p`-DDD rather than op-DDE. (C) 2010 Elsevier Ltd. All rights reserved.

Presumed normal ultrasonographic findings of the testis and epididymis of Botos (Inia Geoffrensis)

Fifteen live adult male botos, or Amazon river dolphins (Inia geoffrensis), were examined using ultrasonography during the yearly capture expedition, between October and November 2005, at the Mamiraua Sustainable Development Reserve, within the Brazilian Amazon (3 degrees S, 65 degrees W). All examinations were performed with a Sonosite 180 plus ultrasound unit in conjunction with a 2- to 5-MHz multifrequency transducer convex array 180 Plus/Elite-C60. Age and maturity estimates were determined considering the body length, weight, and external characteristics. In all examinations, the testes were discerned by the presence of a hyperechoic central line, called the mediastinum testis, a landmark for their identification during ultrasonography. No significant differences in echogenicity were detected on the ultrasonographic appearance of the testes among the studied animals. On adult male botos, apparent parenchymal nodulation of the testis was observed on scanning in most of the animals and probably constituted evidence of reproductive maturity. Using the color Doppler technique, blood flow was detected along the mediastinum testis that progressively decreased toward the periphery of this organ. Little blood flow could be identified by color Doppler. Power Doppler allowed better accuracy to identify testicular vessels, their topography, and their differentiation from adjacent structures. Ultrasonographic examination provides useful data for morphologic characterization of the boto's testes. Examination using Doppler techniques was considered a valuable tool to evidence blood flow through the testicular parenchyma.

Timing of ontogenetic changes of two cranial regions in Sotalia guianensis (Delphinidae)

Despite the fact that heterochronic processes seem to be an important process determining morphological evolution of the delphinid skull, previous workers have not found allometric scaling as relevant factor in the differentiation within the genus Sotalia. Here we analyzed the skull ontogeny of the estuarine dolphin S. guianensis and investigate differential growth and shape changes of two cranial regions the neurocranium and the face in order to evaluate the relevance of cranial compartmentalization on the ontogeny of this structure. Our results show that, even though both cranial regions stop growing at adulthood, the face has higher initial growth rates than the neurocranium. The rate of shape changes is also different for both regions, with the face showing a initially higher, but rapidly decreasing rate of change, while the neurocranium shows a slow decreasing rate, leading to persistent and localized shape changes throughout adult life, a pattern that could be related to epigenetic regional factors. The pattern of ontogenetic shape change described here is similar to those described for other groups of Delphinidae and also match intra and interspecific variation found within the family, suggesting that mosaic heterochrony could be an important factor in the morphological evolution of this group. (C) 2012 Deutsche Gesellschaft fur Saugetierkunde. Published by Elsevier GmbH. All rights reserved.

Gross and microscopic observations on the lingual structure of the franciscana (Pontoporia blainvillei-gervais and d'orbigny, 1844)

In most anatomical studies developed with mammals, the tongue is described as highly differentiated among different species. However, studies on the tongue of aquatic mammals are still limited as compared to those on terrestrial mammals. The aim of this study was to describe the tongue morphology of the Franciscana dolphin (Pontoporia blainvillei) using macroscopic observations, light, and scanning electron microscopy. Microscopically, the dorsal surface was covered by a keratinized stratified epithelium. Salivary gland acini were found on the middle and caudal third of the tongue. The dorsal surface was totally covered by filiform papillae with a connective tissue core and a connective tissue structure round in shape in the middle and caudal regions. Microsc. Res. Tech. 75:737742, 2012. (C) 2012 Wiley Periodicals, Inc.

Feeding habits of Guiana dolphins, Sotalia guianensis, from south-eastern Brazil: new items and a knowledge review

This study presents new information on feeding habits of Guiana dolphins, Sotalia guianensis, in south-eastern Brazil, together with new regression equations to evaluate the weight and length of fish from otoliths, showing an overview on the knowledge about this species' diet in this area. Eighteen stomach contents had been analysed and compared to 180 samples collected in another eight feeding studies. The analysed specimens were either incidentally caught in gillnets used in coastal waters by the fleet based in the Cananeia main harbour (25 degrees 00'S 47 degrees 55'W), south of Sao Paulo State, or found dead in inner waters of the Cananeia estuary between 2003 and 2009. Based on the index of relative importance analysis, the most important fish species were the banded croaker, Paralonchurus brasiliensis. Doryteuthis plei was the most representative cephalopod species. Stellifer rastrifer was the most important fish species observed in dolphins in inner estuarine waters and P. brasiliensis in recovered dolphins from coastal waters. Loliguncula brevis is the only cephalopod species reported from dolphins found in inner estuarine waters up to date. Doryteuthis plei was the most important cephalopod species observed in coastal dolphins. When considering other feeding studies, the most representative fish family in the diet of S. guianensis was Sciaenidae, which is mainly represented by demersal fishes. The main preys of S. guianensis are abundant in the studied areas, which may indicate an opportunistic feeding habit. The majority of them are not the most important target species by the commercial fishery in south-eastern Brazil.

Placentation in dolphins from the Amazon River Basin: the Boto, Inia geoffrensis, and the Tucuxi, Sotalia fluviatilis

A recent reassessment of the phylogenetic affinities of cetaceans makes it timely to compare their placentation with that of the artiodactyls. We studied the placentae of two sympatric species of dolphin from the Amazon River Basin, representing two distinct families. The umbilical cord branched to supply a bilobed allantoic sac. Small blood vessels and smooth muscle bundles were found within the stroma of the cord. Foci of squamous metaplasia occurred in the allanto-amnion and allantochorion. The interhemal membrane of the placenta was of the epitheliochorial type. Two different types of trophoblastic epithelium were seen. Most was of the simple columnar type and indented by fetal capillaries. However, there were also areolar regions with tall columnar trophoblast and these were more sparsely supplied with capillaries. The endometrium was well vascularised and richly supplied with actively secreting glands. These findings are consistent with the current view that Cetacea are nested within Artiodactyla as sister group to the hippopotamids.