The rostral micro-tooth morphology of blue marlin, Makaira nigricans.

Billfish rostra potentially have several functions; however, their role in feeding is unequivocal in some species. Recent work linked morphological variation in rostral micro-teeth to differences in feeding behavior in two billfish species, the striped marlin (Kajikia audax) and the sailfish (Istiophorus platypterus). Here, we present the rostral micro-tooth morphology for a third billfish species, the blue marlin (Makaira nigricans), for which the use of the rostrum in feeding behavior is still undocumented from systematic observations in the wild. We measured the micro-teeth on rostrum tips of blue marlin, striped marlin, and sailfish using a micro-computed tomography approach and compared the tooth morphology among the three species. This was done after an analysis of video-recorded hunting behavior of striped marlin and sailfish revealed that both species strike prey predominantly with the first third of the rostrum, which provided the justification to focus our analysis on the rostrum tips. In blue marlin, intact micro-teeth were longer compared to striped marlin but not to sailfish. Blue marlin had a higher fraction of broken teeth than both striped marlin and sailfish, and broken teeth were distributed more evenly on the rostrum. Micro-tooth regrowth was equally low in both marlin species but higher in sailfish. Based on the differences and similarities in the micro-tooth morphology between the billfish species, we discuss potential feeding-related rostrum use in blue marlin. We put forward the hypothesis that blue marlin might use their rostra in high-speed dashes as observed in striped marlin, rather than in the high-precision rostral strikes described for sailfish, possibly focusing on larger prey organisms.

Spatial Venomics─Cobra Venom System Reveals Spatial Differentiation of Snake Toxins by Mass Spectrometry Imaging.

Among venomous animals, toxic secretions have evolved as biochemical weapons associated with various highly specialized delivery systems on many occasions. Despite extensive research, there is still limited knowledge of the functional biology of most animal toxins, including their venom production and storage, as well as the morphological structures within sophisticated venom producing tissues that might underpin venom modulation. Here, we report on the spatial exploration of a snake venom gland system by matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI), in combination with standard proteotranscriptomic approaches, to enable in situ toxin mapping in spatial intensity maps across a venom gland sourced from the Egyptian cobra (Naja haje). MALDI-MSI toxin visualization on the elapid venom gland reveals a high spatial heterogeneity of different toxin classes at the proteoform level, which may be the result of physiological constraints on venom production and/or storage that reflects the potential for venom modulation under diverse stimuli.

Inner ear sensory system changes as extinct crocodylomorphs transitioned from land to water.

Major evolutionary transitions, in which animals develop new body plans and adapt to dramatically new habitats and lifestyles, have punctuated the history of life. The origin of cetaceans from land-living mammals is among the most famous of these events. Much earlier, during the Mesozoic Era, many reptile groups also moved from land to water, but these transitions are more poorly understood. We use computed tomography to study changes in the inner ear vestibular system, involved in sensing balance and equilibrium, as one of these groups, extinct crocodile relatives called thalattosuchians, transitioned from terrestrial ancestors into pelagic (open ocean) swimmers. We find that the morphology of the vestibular system corresponds to habitat, with pelagic thalattosuchians exhibiting a more compact labyrinth with wider semicircular canal diameters and an enlarged vestibule, reminiscent of modified and miniaturized labyrinths of other marine reptiles and cetaceans. Pelagic thalattosuchians with modified inner ears were the culmination of an evolutionary trend with a long semiaquatic phase, and their pelagic vestibular systems appeared after the first changes to the postcranial skeleton that enhanced their ability to swim. This is strikingly different from cetaceans, which miniaturized their labyrinths soon after entering the water, without a prolonged semiaquatic stage. Thus, thalattosuchians and cetaceans became secondarily aquatic in different ways and at different paces, showing that there are different routes for the same type of transition.

Lacunae rostralis: A new structure on the rostrum of sailfish Istiophorus platypterus.

Recent comparative studies of billfishes (Istiophoridae and Xiphiidae) have provided evidence of differences in the form and function of the rostra (bill) among species. Here, we report the discovery of a new structure, lacuna rostralis, on the rostra of sailfish Istiophorus platypterus, which is absent on the rostra of swordfish Xiphias gladius, striped marlin Kajikia audax and blue marlin Makaira nigricans. The lacunae rostralis are small cavities that contain teeth. They were found on the ventral rostrum surface of all I. platypterus specimens examined and dorsally in half of them. Ventrally, the lacunae rostralis were most prominent in the mid-section of the rostrum. Dorsally, they occurred closer to the tip. The density of lacunae rostralis increased towards the rostrum tip but, because they are smaller in size, the percentage of rostrum coverage decreased. The teeth located within the lacunae rostralis were found to be different in size, location and orientation from the previously identified micro-teeth of billfish. We propose two potential functions of the lacunae rostralis that both relate to the use of the bill in feeding: mechanoreception of prey before tapping it with the bill and more efficient prey handling via the creation of suction, or physical grip.

Savannah roars: The vocal anatomy and the impressive rutting calls of male impala (Aepyceros melampus) - highlighting the acoustic correlates of a mobile larynx.

A retractable larynx and adaptations of the vocal folds in the males of several polygynous ruminants serve for the production of rutting calls that acoustically announce larger than actual body size to both rival males and potential female mates. Here, such features of the vocal tract and of the sound source are documented in another species. We investigated the vocal anatomy and laryngeal mobility including its acoustical effects during the rutting vocal display of free-ranging male impala (Aepyceros melampus melampus) in Namibia. Male impala produced bouts of rutting calls (consisting of oral roars and interspersed explosive nasal snorts) in a low-stretch posture while guarding a rutting territory or harem. For the duration of the roars, male impala retracted the larynx from its high resting position to a low mid-neck position involving an extensible pharynx and a resilient connection between the hyoid apparatus and the larynx. Maximal larynx retraction was 108 mm based on estimates in video single frames. This was in good concordance with 91-mm vocal tract elongation calculated on the basis of differences in formant dispersion between roar portions produced with the larynx still ascended and those produced with maximally retracted larynx. Judged by their morphological traits, the larynx-retracting muscles of male impala are homologous to those of other larynx-retracting ruminants. In contrast, the large and massive vocal keels are evolutionary novelties arising by fusion and linear arrangement of the arytenoid cartilage and the canonical vocal fold. These bulky and histologically complex vocal keels produced a low fundamental frequency of 50 Hz. Impala is another ruminant species in which the males are capable of larynx retraction. In addition, male impala vocal folds are spectacularly specialized compared with domestic bovids, allowing the production of impressive, low-frequency roaring vocalizations as a significant part of their rutting behaviour. Our study expands knowledge on the evolutionary variation of vocal fold morphology in mammals, suggesting that the structure of the mammalian sound source is not always human-like and should be considered in acoustic analysis and modelling.

The remarkable vocal anatomy of the koala (Phascolarctos cinereus): insights into low-frequency sound production in a marsupial species.

Koalas are characterised by a highly unusual vocal anatomy, with a descended larynx and velar vocal folds, allowing them to produce calls at disproportionately low frequencies. Here we use advanced imaging techniques, histological data, classical macroscopic dissection and behavioural observations to provide the first detailed description and interpretation of male and female koala vocal anatomy. We show that both males and females have an elongated pharynx and soft palate, resulting in a permanently descended larynx. In addition, the hyoid apparatus has a human-like configuration in which paired dorsal, resilient ligaments suspend the hyoid apparatus from the skull, while the ventral parts tightly connect to the descended larynx. We also show that koalas can retract the larynx down into the thoracic inlet, facilitated by a dramatic evolutionary transformation of the ventral neck muscles. First, the usual retractors of the larynx and the hyoid have their origins deep in the thorax. Secondly, three hyoid muscles have lost their connection to the hyoid skeleton. Thirdly, the genioglossus and geniohyoid muscles are greatly increased in length. Finally, the digastric, omohyoid and sternohyoid muscles, connected by a common tendinous intersection, form a guiding channel for the dynamic down-and-up movements of the ventral hyoid parts and the larynx. We suggest that these features evolved to accommodate the low resting position of the larynx and assist in its retraction during call production. We also confirm that the edges of the intra-pharyngeal ostium have specialised to form the novel, extra-laryngeal velar vocal folds, which are much larger than the true intra-laryngeal vocal folds in both sexes, but more developed and specialised for low frequency sound production in males than in females. Our findings illustrate that strong selection pressures on acoustic signalling not only lead to the specialisation of existing vocal organs but can also result in the evolution of novel vocal structures in both sexes.

Macrovibrissae and microvibrissae inversion and lateralization in elephants.

Elephants are known for strongly lateralized trunk behaviors, but the mechanisms driving elephant lateralization are poorly understood. Here, we investigate features of elephant mouth organization that presumably promote lateralization. We find the lower jaw of elephants is of narrow width, but is rostrally strongly elongated even beyond the jaw bone. Elephant lip vibrissae become progressively longer rostrally. Thus, elephants have two lateral dense, short microvibrissae arrays and central, less dense long macrovibrissae. This is an inversion of the ancestral mammalian facial vibrissae pattern, where central, dense short microvibrissae are flanked by two lateral macrovibrissae arrays. Elephant microvibrissae have smaller follicles than macrovibrissae. Similar to trunk-tip vibrissae, elephant lip microvibrissae show laterally asymmetric abrasion. Observations on Asian zoo elephants indicate lateralized abrasion results from lateralized feeding. It appears that the ancestral mammalian mouth (upper and lower lips, incisors, frontal microvibrissae) is shaped by oral food apprehension. The elephant mouth organization radically changed, however, because trunk-mediated feeding replaced oral apprehension. Such elephant mouth changes include the upper lip-nose fusion to the trunk, the super-flexible elongated lower jaw, the loss of incisors, and lateral rather than frontal microvibrissae. Elephants' specialization for lateral food insertion is reflected by the reduction in the centering effects of oral food apprehension and lip vibrissae patterns.

Evolution of translational control and the emergence of genes and open reading frames in human and non-human primate hearts.

Evolutionary innovations can be driven by changes in the rates of RNA translation and the emergence of new genes and small open reading frames (sORFs). In this study, we characterized the transcriptional and translational landscape of the hearts of four primate and two rodent species through integrative ribosome and transcriptomic profiling, including adult left ventricle tissues and induced pluripotent stem cell-derived cardiomyocyte cell cultures. We show here that the translational efficiencies of subunits of the mitochondrial oxidative phosphorylation chain complexes IV and V evolved rapidly across mammalian evolution. Moreover, we discovered hundreds of species-specific and lineage-specific genomic innovations that emerged during primate evolution in the heart, including 551 genes, 504 sORFs and 76 evolutionarily conserved genes displaying human-specific cardiac-enriched expression. Overall, our work describes the evolutionary processes and mechanisms that have shaped cardiac transcription and translation in recent primate evolution and sheds light on how these can contribute to cardiac development and disease.

Dynamic three-dimensional computed tomographic imaging facilitates evaluation of the equine cervical articular process joint in motion.

BACKGROUND: Dynamic computed tomography (CT) imaging has been introduced in human orthopaedics and is continuing to gain popularity. With dynamic CT, video sequences of anatomical structures can be evaluated in motion. OBJECTIVES: To investigate the feasibility of dynamic CT for diagnostic imaging of the equine cervical articular process joints (APJs) and to give a detailed description of the APJ movement pattern. STUDY DESIGN: Descriptive cadaver imaging. METHODS: Cervical specimens of twelve Warmblood horses were included. A custom-made motorised testing device was used to position and manipulate the neck specimens and perform dynamic 2D and 3D CT imaging. Images were obtained with a 320-detector-row CT scanner with a 160 mm wide-area (2D) solid-state detector design that allows image acquisition of a volumetric axial length of 160 mm without moving the CT couch. Dynamic videos were acquired and divided into four phases of movement. Three blinded observers used a subjective scale of 1 (excellent) to 4 (poor) to grade the overall image quality in each phases of motion cycle. RESULTS: With an overall median score of 1 the image quality, a significantly lower score was observed in the dynamic 3D videos over the four phases by the three observers compared with the 2D videos for both flexion (3D 95% CI: 1-2 and 2D 95% CI: 1-3; P = .007) and extension movement (3D 95% CI: 1-2 and 2D 95% CI: 1-3; P = .008). Median Translational displacement of the APJ surface was significantly greater in flexion than in extension movement (P = .002). MAIN LIMITATIONS: The small number of specimens included. Excision of spines and removal of musculature. CONCLUSIONS: The study is a first step in the investigation of the potential of dynamic 3D CT in veterinary medicine, a technique that has only begun to be explored and leaves much room for refinement prior to its introduction in routine practice. CT with a detector coverage of 16 cm and a rotation speed of 0.32 seconds provides high-quality images of moving objects and gives new insight into the movement pattern of equine cervical APJs.

The functional anatomy of elephant trunk whiskers.

Behavior and innervation suggest a high tactile sensitivity of elephant trunks. To clarify the tactile trunk periphery we studied whiskers with the following findings. Whisker density is high at the trunk tip and African savanna elephants have more trunk tip whiskers than Asian elephants. Adult elephants show striking lateralized whisker abrasion caused by lateralized trunk behavior. Elephant whiskers are thick and show little tapering. Whisker follicles are large, lack a ring sinus and their organization varies across the trunk. Follicles are innervated by ~90 axons from multiple nerves. Because elephants don't whisk, trunk movements determine whisker contacts. Whisker-arrays on the ventral trunk-ridge contact objects balanced on the ventral trunk. Trunk whiskers differ from the mobile, thin and tapered facial whiskers that sample peri-rostrum space symmetrically in many mammals. We suggest their distinctive features-being thick, non-tapered, lateralized and arranged in specific high-density arrays-evolved along with the manipulative capacities of the trunk.

Trigeminal ganglion and sensory nerves suggest tactile specialization of elephants.

Sensory nerves are information bottlenecks giving rise to distinct sensory worlds across animal species.1 Here, we investigate trigeminal ganglion2,3 and sensory nerves4 of elephants. The elephant trigeminal ganglion is very large. Its maxillary branch, which gives rise to the infraorbital nerve innervating the trunk, has a larger diameter than the animal's spinal cord, i.e., trunk innervation is more substantive than connections of the brain to the rest of the body. Hundreds of satellite cells surround each trigeminal neuron, an indication of exceptional glial support to these large projection neurons.5-7 Fiber counts of Asian elephant infraorbital nerves of averaged 4,00,000 axons. The infraorbital nerve consists of axons that are ∼10 µm thick and it has a large diameter of 17 mm, roughly 3 times as thick as the optic and 6 times as thick as the vestibulocochlear nerve. In most mammals (including tactile specialists) optic nerve fibers8-10 greatly outnumber infraorbital nerve fibers,11,12 but in elephants the infraorbital nerve fiber count is only slightly lower than the optic nerve fiber count. Trunk innervation (nerves and ganglia) weighs ∼1.5 kg in elephant cows. Our findings characterize the elephant trigeminal ganglion as one of the largest known primary sensory structures and point to a high degree of tactile specialization in elephants.

Genomic basis for skin phenotype and cold adaptation in the extinct Steller's sea cow.

Steller's sea cow, an extinct sirenian and one of the largest Quaternary mammals, was described by Georg Steller in 1741 and eradicated by humans within 27 years. Here, we complement Steller's descriptions with paleogenomic data from 12 individuals. We identified convergent evolution between Steller's sea cow and cetaceans but not extant sirenians, suggesting a role of several genes in adaptation to cold aquatic (or marine) environments. Among these are inactivations of lipoxygenase genes, which in humans and mouse models cause ichthyosis, a skin disease characterized by a thick, hyperkeratotic epidermis that recapitulates Steller's sea cows' reportedly bark-like skin. We also found that Steller's sea cows' abundance was continuously declining for tens of thousands of years before their description, implying that environmental changes also contributed to their extinction.

Use of deep learning for structural analysis of computer tomography images of soil samples.

Soil samples from several European countries were scanned using medical computer tomography (CT) device and are now available as CT images. The analysis of these samples was carried out using deep learning methods. For this purpose, a VGG16 network was trained with the CT images (X). For the annotation (y) a new method for automated annotation, 'surrogate' learning, was introduced. The generated neural networks (NNs) were subjected to a detailed analysis. Among other things, transfer learning was used to check whether the NN can also be trained to other y-values. Visually, the NN was verified using a gradient-based class activation mapping (grad-CAM) algorithm. These analyses showed that the NN was able to generalize, i.e. to capture the spatial structure of the soil sample. Possible applications of the models are discussed.

Re-Emergence and Spread of Haemorrhagic Septicaemia in Germany: The Wolf as a Vector?

Since 2010, outbreaks of haemorrhagic septicaemia (HS) caused by Pasteurella (P.) multocida capsular type B (PmB) emerged in Germany. In 2017, we noticed a close spatiotemporal relationship between HS outbreak sites and wolf (Canis lupus) territories. Thus, the main objectives of our study were to investigate the molecular epidemiology of German PmB-HS-isolates and to assess the role of wolves as putative vectors of this pathogen. We collected 83 PmB isolates from HS outbreaks that occurred between 2010 and 2019 and sampled 150 wolves, which were found dead in the years 2017 to 2019, revealing another three PmB isolates. A maximum-likelihood-based phylogeny of the core genomes of 65 PmB-HS-isolates and the three PmB-wolf-isolates showed high relatedness. Furthermore, all belonged to capsular:LPS:MLST genotype B:L2:ST122RIRDC and showed highly similar virulence gene profiles, but clustered separately from 35 global ST122RIRDC strains. Our data revealed that German HS outbreaks were caused by a distinct genomic lineage of PmB-ST122 strains, hinting towards an independent, ongoing epidemiologic event. We demonstrated for the first time, that carnivores, i.e., wolves, might harbour PmB as a part of their oropharyngeal microbiota. Furthermore, the results of our study imply that wolves can carry the pathogen over long distances, indicating a major role of that animal species in the ongoing epidemiological event of HS in Germany.

Chronic heart failure due to severe endocardiosis in a Gambian giant pouched rat (Cricetomys gambianus).

A 1.5-yr-old captive male Gambian giant pouched rat (Cricetomys gambianus) died after suffering from anorexia, weakness, and dyspnea for 3 wk. Thoracic radiographs of thorax and abdomen and computed tomography showed a severe biventricular enlargement of the heart and a moderate hepatomegaly. Necropsy revealed a severe, bilateral hypertrophic cardiomyopathy and dilation of the right ventricle due to multifocal bilateral, valvular endocardiosis of all atrioventricular valves and acute hepatic congestion. Histologically, the atrioventricular valves were multifocally thickened by a marked endocardiosis with stromal accumulation of Periodic acid-Schiff (PAS)-positive, mucinous material. Although common in dogs, endocardiosis has not been described in Nesomyidaes. As in other affected species, the pathogenesis of endocardiosis in this species remains unclear.

Computed Tomographic Evaluation of Adjacent Segment Motion after Ex Vivo Fusion of Equine Third and Fourth Cervical Vertebrae.

OBJECTIVE: Surgical fusion of vertebral segments is a treatment option for horses with cervical stenotic myelopathy or cervical fracture.Degenerative disease affecting adjacent vertebral segments is a reported complication following surgical vertebral fusion in other species, termed adjacent segment disease. The aim of this study was to evaluate the impact of cervical vertebral fusion on the biomechanics of adjacent vertebral segments in the horse. STUDY DESIGN: Neck specimens of 12 horses were assessed using computed tomographic imaging. Range of motion (ROM) was determined by measuring the maximum sagittal flexion, extension and lateral bending between C2 and C5. C3/4 was subsequently fused using a standard locking compression plate and locking head screws and computed tomographic scans and ROM measurements were repeated. RESULTS: Prior to intervertebral fusion, a significant increase in ROM along the vertebral segments from cranial to caudal was observed. Range of motion measurements of C3/4 decreased significantly after fusion (p = 0.01).Range of motion of the adjacent segments (C2/3 and C4/5) did not change significantly after fusion. CONCLUSION: Fusion of one cervical intervertebral joint did not affect the ROM of the adjacent vertebral segments. Further research investigating the implications of vertebral fusion on the intervertebral pressure in the equine patient is indicated.

Paraplegia in a Bornean orangutan (Pongo pygmaeus pygmaeus) due to multiple myeloma.

BACKGROUND: A 38-year-old male Bornean orangutan (Pongo pygmaeus pygmaeus) developed progressive hind leg paresis. A computed tomography scan of the vertebral column revealed soft tissue type densities within vertebral bones. METHODS AND RESULTS: At necropsy infiltrating tumor masses were found in the vertebral bodies, protruding into the spinal canal and compressing the spinal cord. Microscopically neoplastic plasma cells infiltrated the vertebral bodies and adjacent soft tissues. Immunohistochemically, tumor cells tested positive for B cell markers (CD38, CD79alpha), kappa, and lambda light chains, while vimentin, GFAP, S100, and CD138 were not expressed. The tumor was classified as multiple myeloma on the basis of radiographic, pathological, and immunohistochemical findings. CONCLUSIONS: This first systematic case description on multiple myeloma in a non-human primate revealed many similarities with the disease in humans and the immunohistochemical tools proved suitable for their use in the orangutan.

Optimized DNA sampling of ancient bones using Computed Tomography scans.

The prevalence of contaminant microbial DNA in ancient bone samples represents the principal limiting factor for palaeogenomic studies, as it may comprise more than 99% of DNA molecules obtained. Efforts to exclude or reduce this contaminant fraction have been numerous but also variable in their success. Here, we present a simple but highly effective method to increase the relative proportion of endogenous molecules obtained from ancient bones. Using computed tomography (CT) scanning, we identify the densest region of a bone as optimal for sampling. This approach accurately identifies the densest internal regions of petrous bones, which are known to be a source of high-purity ancient DNA. For ancient long bones, CT scans reveal a high-density outermost layer, which has been routinely removed and discarded prior to DNA extraction. For almost all long bones investigated, we find that targeted sampling of this outermost layer provides an increase in endogenous DNA content over that obtained from softer, trabecular bone. This targeted sampling can produce as much as 50-fold increase in the proportion of endogenous DNA, providing a directly proportional reduction in sequencing costs for shotgun sequencing experiments. The observed increases in endogenous DNA proportion are not associated with any reduction in absolute endogenous molecule recovery. Although sampling the outermost layer can result in higher levels of human contamination, some bones were found to have more contamination associated with the internal bone structures. Our method is highly consistent, reproducible and applicable across a wide range of bone types, ages and species. We predict that this discovery will greatly extend the potential to study ancient populations and species in the genomics era.

Progressive ataxia in a captive North American river otter (Lontra canadensis) associated with brain stem spheroid formation.

Progressive ataxia, particularly of the hind limbs, developed in a paretic female juvenile captive North American river otter (Lontra canadensis). Severe bilaterally symmetrical axonal degeneration in the medulla oblongata (nucleus gracilis and nucleus cuneatus) and multiple scattered spheroids in the region of the pons were detected microscopically. This is the first description of ataxia associated with axonal degeneration in otters.

Surgical approach for tentorial meningiomas in cats: a review of six cases.

The surgical technique for removal of tentorial meningiomas is described on six cats using a unilateral temporal supracerebellar transtentorial approach. Complete gross tumour resection was achieved in four of six cats. In one cat, only subtotal resection was achieved. One cat died shortly after surgery because of extensive cerebral haemorrhage. The surgical approach, combined with cisternal or ventricular cerebrospinal fluid puncture and an open-window technique (tumour fenestration and enucleation) provided sufficient visibility and tumour accessibility without excessive manipulation of the brain parenchyma. In all patients, a postoperative transient worsening of the clinical signs was observed. The neurological signs resolved with time with the exception of blindness in two cats. All five surviving cats were monitored for a mean follow-up time of 19 months (median 20 months; range 6-30 months). All patients died or were euthanased because of tumour regrowth within the follow-up period. Although challenging, surgical treatment is a useful therapeutic measure in the treatment of cats presenting with tentorial meningiomas.