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Rapid growth in aquaculture has resulted in high-density production systems in ecologically and geographically novel conditions in which the emergence of diseases is inevitable. Well-characterized methods for detection and surveillance of infectious diseases are vital for rapid identification, response, and recovery to protect economic and food security. We implemented a proof-of-concept approach for virus detection using a known high-consequence fish pathogen, infectious salmon anemia virus (ISAV), as the archetypal pathogen. In fish infected with ISAV, we integrated histopathology, virus isolation, whole-genome sequencing (WGS), electron microscopy (EM), in situ hybridization (ISH), and reverse transcription real-time PCR (RT-rtPCR). Fresh-frozen and formalin-fixed tissues were collected from virus-infected, control, and sham-infected Atlantic salmon ( Salmo salar ). Microscopic differences were not evident between uninfected and infected fish. Viral cytopathic effect was observed in cell cultures inoculated with fresh-frozen tissue homogenates from 3 of 3 ISAV-infected and 0 of 4 uninfected or sham-infected fish. The ISAV genome was detected by shotgun metagenomics in RNA extracted from the medium from 3 of 3 inoculated cell cultures, 3 of 3 infected fish, and 0 of 4 uninfected or sham-infected fish, yielding sufficient coverage for de novo assembly. An ISH probe against ISAV revealed ISAV genome in multiple organs, with abundance in renal hematopoietic tissue. Virus was detected by RT-rtPCR in gill, heart, kidney, liver, and spleen. EM and metagenomic WGS from tissues were challenging and unsuccessful. Our proof-of-concept methodology has promise for detection and characterization of unknown aquatic pathogens and also highlights some associated methodology challenges that require additional investigation., Competing Interests: Declaration of conflicting interestsThe authors declared no potential conflicts of interest with respect to research, authorship, and/or publication of this article.

Lipofuscin accumulation has been observed in human coronary arteries but whether or not myocardial tissue can release lipofuscin generated within cardiomyocytes must be clarified, as this may provide indicators for future anti-ageing research. The hearts of Sprague Dawley rats, aged 6-24 months, were embedded in resin and ultrathin sections cut for electron microscopy. Lipofuscin granules were abundant in cardiomyocytes. Cardiomyocytes were seen to release lipofuscin granules into the myocardial interstitium as cytoplasmic fragments with irregular protrusions on the sarcolemma surface. The cytoplasmic fragments entering the stroma fused directly with the endothelial cells of adjacent capillaries, delivering lipofuscin to the vessel wall. These fragments were also seen to be engulfed by stromal macrophages or fused with fibroblasts, which then combined with capillary endothelial cells to deliver lipofuscin to the vessel wall. Some cytoplasmic fragments disaggregated and formed membrane-like waste, which travelled to the vessel wall from the myocardial stroma as soluble fine particles via diffusion or pinocytosis of capillary endothelial cells. Lipofuscin entered the vascular wall and endothelial cells, forming large and small protrusions or folds that transported the lipofuscin to the vascular lumen and bloodstream., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

The current study aims to present differences between the pecten oculi of different avian species through morphologic, macroscopic, light, and electron microscopic examinations. The study is a comprehensive research on seven avian species (sparrowhawk, hawk, magpie, swan, heron, pheasant, duck). The right eyes of the animals utilized in the study were removed for light microscopic examination, whereas their left eyes were removed for electron microscopic and macroscopic examinations. Morphometric analyses, as well as stereo and light microscopic measurements, were carried out on the pecten oculi of the animals. Given all these data, it was determined that the height of the pecten oculi did not differ among the species in the study; however, the pecten oculi were larger in birds with the highest value compared to the other species in the macroscopic measurements. Also, the pecten oculi vessels were larger, and the number of melanocytes was higher in keen eyesight, raptor, and migratory birds with large bulbus oculi. All these data suggest that the pecten oculi not only supplies nutrient to the retina but also contributes to sharp vision during migration and hunting, UV absorption from sunlight, as well as preservation of intraocular equilibrium., (© 2023 Wiley-VCH GmbH. Published by John Wiley & Sons Ltd.)

Aims: To investigate the pathogenesis of a disease in takahē (Porphyrio hochstetteri) with intracytoplasmic inclusion bodies in lower motor neurons., Methods: Four birds aged between 5 and 12 years, from three different wildlife sanctuaries in New Zealand were examined. Of these, only one had signs of spinal dysfunction in the form of paresis. Stained paraffin sections of tissues were examined by light microscopy and immunostained sections of the ventral horn of the spinal cord by confocal microscopy. Epoxy resin sections of the spinal cord from the bird with spinal dysfunction were examined by electron microscopy., Results: Two types of inclusion bodies were noted, but only in motor neurons of the ventral spinal cord and brain stem. These were large globoid eosinophilic bodies up to 5 µm in diameter, and yellow/brown granular inclusions mostly at the pole of the cell. The globoid bodies stained with Luxol fast blue but not with periodic acid Schiff (PAS), or Sudan black. The granular inclusions stained with Luxol fast blue, PAS and Sudan black. Both bodies were slightly autofluorescent. On electron microscopy the globoid bodies had an even electron-dense texture and were bound by a membrane. Beneath the membrane were large numbers of small intraluminal vesicles. The smaller granular bodies were more heterogeneous, irregularly rounded and membrane-bound accumulations of granular electron-dense material, often with electron-lucent vacuoles. Others were more vesicular but contained varying amounts of electron-dense material. The large globoid bodies did not immunostain for lysosomal markers lysosomal associated protein 1 (LAMP1) or cathepsin D, so were not lysosomal. The small granular bodies stained for cathepsin D by a chromogenic method.A kindred matrix analysis showed two cases to be as closely related as first cousins, and another case was almost as closely related to one of them, but the fourth bird was unrelated to any other., Conclusions: It was concluded that this was an endoplasmic reticulum storage disease due to a specific protein misfolding within endoplasmic reticulum. It was rationalised that the two types of inclusions reflected the same aetiology, but that misfolded protein in the smaller granular bodies had entered the lysosomal system via endoplasmic reticulum autophagy. Although the cause was unclear, it most likely had a genetic aetiology or predisposition and, as such, has clinical relevance.

We document the frequency and morphological and immunohistochemical features of inclusion bodies in uterine smooth muscle cells in 56 (76%) of 74 investigated pet rabbits (Oryctolagus cuniculus). Inclusion bodies began to appear at the age of 2 years and their frequency increased with age (P = 0.047, r = 0.33). They ranged from 5 to 20 μm in diameter, were slightly basophilic to amphophilic with well-delimited oval bodies in haematoxylin and eosin-stained tissue sections and formed in the cytoplasm of the uterine smooth muscle cells with displacement of the cell nuclei. The inclusion bodies were positive with periodic acid-Schiff, Best's carmine, Lugol's iodine and Grocott's methenamine silver methods. They were immunoreactive to a monoclonal antibody raised against human polyglucosan and negative with monoclonal antibodies for several intermediate filament proteins. Electron microscopy revealed that they were non-membranous structures composed of electron-dense amorphous material. The morphological, histochemical, immunohistochemical and ultrastructural features of the inclusion bodies in the rabbi uteri were similar to those of human polyglucosan bodies (PGBs). PGBs appear to occur at a high frequency in the uterus of rabbits, which are known to be susceptible to uterine diseases., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

Background: The epiphysis cerebri (pineal gland) is a small-sized, photo neuroendocrine organ in the brain of most vertebrates. Their effect is through secretion of melatonin, a serotonin-derived hormone which is stimulated by darkness and inhibited by light and modulates the circadian rhythm; light and dark cycle like a biological clock, sleep patterns (sleep-wake cycle), and sexual development., Aim: This study aimed to identify and differentiate the different cell types filling the pineal gland parenchyma of mature male sheep., Methods: Pineal glands were collected and sliced parasagitally then processed histologically for light and electron microscopic examinations., Results: Two main cell types; pinealocytes and astrocytes were recognized within the gland parenchyma. Pinealocytes were the chief parenchymatus cells that occupied the largest volume of the gland and were classified according to the nuclear pictures (activity status) into two subtypes; pinealocytes I (pale subtype, active) and II (dark subtype, inactive). Astrocyte neuroglial cells had cytoplasmic processes which form a huge supportive framework between the pinealocytes and clarified two types; type I were elongated cells with elongated snake shaped nucleus and type II were smaller in size, with oval nuclei. Another marginal cell type was identified as a neuron-like cell which appeared larger in size than others and distributed sporadically, has eccentric oval nucleus with prominent nucleoli and single, long cytoplasmic process that branched at its terminal forming T-shaped process looks like pseudo unipolar neuron. Moreover, aggregations of pigment granules were markedly observed in the intercellular spaces and also near the blood capillaries. With transmission electron microscope (TEM) a special characteristic feature of pinealocytes; synaptic ribbons were recognized that appeared as bands of electron-dense material with several synaptic spherules; vesicles adjacent to its surface helping in the multivesicular release., Conclusion: The gland parenchyma revealed two main cell types; pinealocytes and astrocytes. Each one was subdivided into two subtypes; I and II. The first one was classified according to their nuclear pictures (activity status) and the second one was according to their shape, size, and cytoplasmic processes. Other cell types were also identified as neuronal and pigmented-like cells in the pineal matrix., Competing Interests: The authors declare that there is no conflict of interest.

Plasmodium falciparum remains one of the world's deadliest diseases and with ongoing concerns of evolving drug resistance, there is a need for continued refinement of the Plasmodium coatneyi infection model in macaques to study severe malaria. As such, the systemic ultrastructural lesions associated with P. coatneyi infection in splenectomized rhesus macaques was evaluated in 6 animals. Autopsy samples from multiple areas of the central nervous system (CNS), kidneys, heart, liver, and lungs of all 6 animals were processed for electron microscopy. A systematic analysis of the ultrastructural changes associated with the plasmodium was undertaken by multiple pathologists to ensure consensus. All tissues exhibited marked sequestration of infected red blood cells comprised either of cytoadherence to endothelium or rosette formation, associated with variable degrees of host cell damage in a range of tissues that in severe cases resulted in necrosis. This is the first complete systemic evaluation of ultrastructural tissue lesions in P. coatneyi -infected rhesus macaques, and the findings have important implications evaluating of the use of this model for the study of severe malaria caused by P. falciparum in humans.

A 13-year-old female Miniature Dachshund showed seizures and an intracranial mass was seen in the left temporal lobe. Three months after first surgical resection, a recurrence lesion was recognized. Histopathologically, proliferation of spindle cells with diffuse rosette-like eosinophilic deposits, which stained blue with Masson's trichrome stain, was observed. In electron microscopy, the rosette-like deposits were consisted of bundles of minute filaments which were assumed to be collagenous fibrils. Immunohistochemically, the neoplastic cells were positive for vimentin and cytokeratin, and negative for E-cadherin, S100, glial fibrillary acidic protein (GFAP), oligodendrocyte transcription factor (Olig2), and CD204. The rosette-like collagen deposits were positive for type 4 collagen and negative for type 1 collagen. In this report, we describe histopathological features of a canine meningioma with the rosette-like collagen deposits.

Ultimobranchial body (UBB) remnant was considered as an enigma till the last few years, then it was recognized as a necessary organ where it is the origin of the parafollicular cells. The samples were fixed and processed for the histological and electron microscopic examination. Macroscopically, the UBB remnant appeared as a white mass at the end of the cranial one-third of the thyroid lobe. It was composed of solid cell nest, cluster of cells and small thyroid follicles. Transmission electron microscope showed some round cells containing euchromatic nuclei, numerous parafollicular cells with darkly stained granules and paler ones. It also showed some mast cells with heterochromatic nuclei and large darkly stained granules. The parafollicular cells were distributed throughout the thyroid gland but concentrated within the UBB remnant. To our knowledge, our findings represent very unique histological manifestations specially the ultra-structural ones which revealed an original finding about the new clear type of cells suggested to be a UBB remnant and ensure those of the light microscope., (© 2022 Wiley-VCH GmbH.)

The morphological characteristics of the yellow-legged gull's photoreceptors and cone distribution were studied using light and electron microscopy. In wholemount fresh retinas, five different coloured oil droplets located in the cone inner segments could be seen and characterized by colour, diameter and stratification. The photoreceptors were classified by comparing the fresh and fixed vertical sections under a light and electron microscope. Rods were easily distinguished from cones based on the outer segment morphology and the absence of oil droplets in their inner segments. Four types of single cones were associated with red, yellow, colourless and transparent oil droplets. Unequal double cones comprised a long principal member with a green oil droplet and an accessory short member containing a green microdroplet which was highly electron-dense under electron microscopy. The different types of oil droplets were counted from microphotographs of fresh retinal samples in 20 regions. The density, percentage and diameter of the oil droplets were determined. The results showed that central regions had the highest oil droplet density which decreased towards the retinal periphery in all quadrants. Moreover, the oil droplet density was higher in the dorsotemporal quadrant than in other retinal regions. The average density of the red oil droplets was highest in the central areas, whereas colourless oil droplets had the highest density throughout the retina. In contrast, transparent oil droplets had the lowest density across all the regions of the retina. Finally, the retinal resolution was 52.61 cycles/degree. It was calculated using the posterior nodal distance and the oil droplet diameter. The work concludes by discussing the significance of the relative proportion of different cone types across the retina., (© 2021 The Authors. Anatomia, Histologia, Embryologia published by Wiley-VCH GmbH.)

Morphological development of the lamina epithelialis mucosae (LEM) of the ileum of broiler chicken was evaluated using light and transmission electron microscopic techniques. Ileal sections were collected on embryonic days (ED) 14, 17 and 19, as well as post-hatch days (PD) 1, 3, 5, 7, 14, 21, 28, 35, 42 and 56. The result showed that the ileal LEM, which were poorly defined at embryonic days 14 and 17, consisted of enterocytes and some atypically shaped goblet cells. Apico-lateral tight junctions and irregularly shaped microvilli were associated with the enterocytes at ED 14 and 17. The enterocyte microvilli were more uniform in shape and distribution at ED 19. The embryonic goblet cells were varied in shape and possessed basally displaced, star-shaped nuclei and small apical cytoplasmic vacuoles. During the post-hatch ages, the LEM was a typical epithelium with a single layer of columnar-shaped enterocytes that became highly elongated at post-hatch day (PD) 7. The goblet cells were characteristically 'wine-glass' shaped. Follicle-associated epithelium (FAE) showing numerous lymphocytes among the enterocytes occurred in the post-hatch LEM. The intra-epithelial lymphocytes (IEL) were first encountered at PD 1, but increased several folds within the first two weeks post-hatch. Entero-endocrine cells were observed in the epithelium from PD 21. Finally, from this study, it is obvious that enterocytes and small atypically shaped goblet cells occur in the ileal LEM during the pre-hatch period, but these cells assume adult morphological characteristics after hatch. Thus, the cells of the ileal LEM play strategic absorptive, secretory and protective roles in the gut., (© 2021 Wiley-VCH GmbH.)

Aquaculture is an increasingly important food resource, but its sustainability is often limited by disease. In Scombridae fishes, puffy snout syndrome (PSS) is a debilitating condition where tumor-like collagenous growths form around the eyes, nares, and mandibles which impair vision and feeding and frequently lead to mortality. While PSS is considered an infectious or metabolic disease, no disease agents or promoters have been identified. Here, we used electron microscopy (EM) to describe the cellular pathology and search for etiological agents of PSS in Pacific mackerel Scomber japonicus, the first use of this approach for PSS. We examined aquaculture specimens across a range of apparent PSS severity, comparing the results to both wild and aquaculture asymptomatic mackerel. EM imagery consistently revealed viral-like particles in PSS samples, as well as the uniform absence of bacteria, protists, fungi, and other multicellular parasites. In addition to viral-like particles, symptomatic fish had a higher mean percentage of swollen and disintegrating mitochondria than both asymptomatic aquaculture and wild mackerel. This suggests that degraded mitochondria may be related to PSS and could be important to further understanding the origin, promoters, and prevention of PSS. This study serves as a first step in identifying the etiological agents of PSS.

This study investigated the histomorphological, ultrastructural and morphometrical postnatal developmental changes in the rabbit fundic region, especially during changing of the feeding intake. Seventy-two New Zealand rabbits (V-Line breed) at the ages of 1, 7, 15, 23, 30 and 60 days were obtained for light and electron microscopy and morphometric studies of the fundic region. The newborn rabbit's fundic wall was thin and organized into mucosa, submucosa, musclosa and serosa, with a significant increase in thickness with ageing. The fundic glands were few at the first week of life, then increased in length and diameter compared to the preceding age with prominent zonation at 23 days. The gastric pits appeared wide and deep at the first week of life then became typically narrow and shallow at the third week. The mucous cells were the main cell types lining the fundic glands in the first week of life. These cells showed remoulding with a marked increase in Periodic Acid-Schiff reactivity with age. Parietal cells were differentiated earlier (on the first day of life) than the chief cells and distributed at the neck and basal zones. Chief cells differentiated at 15 days old at the base of the glands, followed by an increase in the number and activity. Few active enteroendocrine cells were first seen at 15 days old and then widely distributed throughout the glands. Conclusion: Pronounced histomorphological changes in the fundic mucosal layer, especially the surface and glandular epithelium, correlate with the postnatal rabbit-feeding intake changes., (© 2021 Wiley-VCH GmbH.)

Background: The cellular mechanisms involved in the development of proximal tubules are not only associated with morphogenesis in fetal life, but also with restoration of damaged tubules in adulthood. Knowledge about morphological features of cell differentiation and proliferation along the developing tubule is insufficient, which hinders identification of the cellular origin., Objectives: This study aimed to investigate ultrastructures of the proximal tubule at different stages of nephrogenesis., Methods: Electron microscopy was used and guided by computer-assisted tubular tracing to identify the cellular structures., Results: Renal vesicles and S-shaped bodies revealed more proliferative features, such as densely-packed fusiform-shaped cells with numerous protein-producing organelles than membrane specializations typical for mature tubules. At the capillary-loop stage the proximal tubules demonstrated all characteristics of the mature tubules, but not as developed, including shorter but densely packed microvilli, fewer lateral processes with cell-cell contacts, lower basal membrane infoldings, and lower mitochondrial volume density. However, they exhibited an elaborated endocytic system above the nucleus, indicating a membrane transport is being established. Abundant free- and endoplasmic reticulum-adhered ribosomes and Golgi complexes reflected active protein synthesis for cell growth and proliferation. Interestingly, electron dense cells were occasionally intermixed with electron lucent cells characterized by various organelles in less cytosol and a larger nucleus with abundant euchromatin, which is a feature of active proliferation., Conclusions: These ultrastructures indicate that the morphogenesis of the developing proximal tubule corresponds to the gradually established physiological activities. The two different cellular electron densities may suggest distinctive differentiation of the cells along the tubule., (© 2021 The Authors. Veterinary Medicine and Science published by John Wiley & Sons Ltd.)

Cutaneous warts are the common clinical feature of infection with Bovine papillomavirus (BPV), and it is commonly known as bovine papillomatosis. It causes significant economic losses, especially in the dairy sector. The aim of this study was surveillance of the circulating strains of BPV in four Egyptian governorates and characterization by electron microscopy. Warts skin lesions and whole blood from seventy-eight native breed cattle were obtained. Molecular detection using two different sets of primers, phylogenetic analysis, and electron microscopy were carried out. The obtained results showed that using FAP59/FAP64 primer set is more sensitive than the MY09/My11 primer set in the detection of the papilloma L1 gene either in the blood or in the skin lesion. Sequence analysis of the partially amplified L1 gene revealed 4 different strains belonging to Deltapapillomavirus 4. Only Alfayoum_Deltapapillomavirus_2018 (accession no: MW018705) was found to be closely related to the strain previously isolated in different Egyptian governorates in 2017, and 2 strains were closely related to an isolate of equine origin. Electron microscopy examination of the skin lesions showed the presence of negatively stained rounded, non-enveloped virus particles with a size of 60 nm in diameter. In conclusion, continuous surveillance and characterization of the circulating strains using multiple sets of primers are important. Efficient biosecurity measures must be applied to decrease transmission of papillomavirus between the different animal species, especially in the mixed management system.

The aim of this study was to establish a methodology of cryopreservation of cattle oocytes and the quality assessment of oocytes and subsequent embryos produced in vitro under our laboratory conditions. Previously in vitro matured (IVM) oocytes were vitrified in minimum volume by ultra-rapid cooling technique. The oocytes were put into the equilibration solution (3% ethylene glycol in M199-HEPES + 10% foetal bovine serum) for 12 min, transferred to vitrification solution (30% ethylene glycol + 1 M sucrose in M199-HEPES + 10% foetal bovine serum) at room temperature for 25 s, then placed onto nickel electron microscopy grids and plunged into liquid nitrogen. After warming 75% of the oocytes were assessed as viable. Part of viable oocytes was taken for electron microscopy, the remaining oocytes were fertilized in vitro, and the presumptive zygotes were cultured until the blastocyst stage. Embryo cleavage and blastocyst rates in vitrified group after warming were 64.98% and 17.3%, resp. versus 70.72% and 25.54% in the control group (P < 0.05). No significant differences were found in the blastocyst total cell number, TUNEL and dead cell indexes between both groups. Ultrastructure of vitrified oocytes showed damages in smooth endoplasmic reticulum (SER) vesicles and lipid droplets as well as irregular arrangement of solitary cortical granules. Several mitochondria were damaged and the microtubules around the chromosomes were less occurred compared to the control group. However, the extent of injuries was lower than reported by other authors studying the ultrastructure of vitrified bovine oocytes, what is also supported by the better development of our oocytes after IVF. In conclusion, the designed oocyte vitrification technique ensures obtaining the blastocysts of the quality comparable to the fresh oocytes., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Porcine sapelovirus (PSV) is a causative agent of acute diarrhoea, pneumonia and reproductive disorders in swine. Since PSV infection interrupts the growth of other viruses due to its high replication capability in cell culture, the prevention of PSV replication is a keystone to the isolation of non-PSV agents from PSV-contaminated samples. In the present study, we established the PSV infection-resistant cell line N1380 and isolated three mammalian orthoreoviruses (MRV) strains, sR1521, sR1677 and sR1590, from swine in Taiwan. These Taiwanese isolates induced an extensive cytopathic effect in N1380 cells upon infection. The complete and empty virus particles were purified from the cell culture supernatants. Next-generation sequencing analyses revealed that the complete virus particles contained 10 segments, including 3 large (L1, L2 and L3), 3 medium (M1, M2 and M3) and 4 small (S1, S2, S3 and S4) segments. In contrast, the empty virus particles without genome were non-infectious. Phylogenetic analyses revealed that the Taiwanese strains belong to serotype 2 MRV (MRV2). We established an ELISA for the detection of IgG antibody against MRV2 by using the empty virus particles as the antigen. A total of 540 swine and 95 wild boar serum samples were collected in Japan, and the positive rates were 100% and 52.6%, respectively. These results demonstrated that MRV infection occurred frequently in both swine and wild boar in Japan. We established a cell line that is efficient for the isolation of MRV, and the ELISA based on the naturally occurring empty particles would be of great value for the surveillance of MRV-related diseases., (© 2020 Blackwell Verlag GmbH.)

Background: Ovine pulmonary adenocarcinoma (OPA) is a neoplastic disease caused by exogenous Jaagsiekte Sheep Retrovirus (exJSRV). The prevalence of JSRV-related OPA in Eastern European countries, including Romania is unknown. We aimed to investigate: the prevalence and morphological features of OPA (classical and atypical forms) in the Transylvania region (Romania), the immunophenotype of the pulmonary tumors and their relationships with exJSRV infection. A total of 2693 adult ewes slaughtered between 2017 and 2019 in two private slaughterhouses from Transylvania region (Romania) was evaluated. Lung tumors were subsequently assessed by cytology, histology, immunocytochemistry, immunohistochemistry, electron microscopy and DNA testing., Results: Out of 2693 examined sheep, 34 had OPA (1.26% prevalence). The diaphragmatic lobes were the most affected. Grossly, the classical OPA was identified in 88.24% of investigated cases and the atypical OPA in 11.76% that included solitary myxomatous nodules. Histopathology results confirmed the presence of OPA in all suspected cases, which were classified into acinar and papillary types. Myxoid growths (MGs) were diagnosed in 6 classical OPA cases and in 2 cases of atypical form. Lung adenocarcinoma was positive for MCK and TTF-1, and MGs showed immunoreaction for Vimentin, Desmin and SMA; Ki67 expression of classical OPA was higher than atypical OPA and MGs. JSRV-MA was identified by IHC (94.11%) in both epithelial and mesenchymal cells of OPA. Immunocytochemistry and electron microscopy also confirmed the JSRV within the neoplastic cells. ExJSRV was identified by PCR in 97.05% of analyzed samples. Phylogenetic analysis revealed the presence of the exJSRV type 2 (MT809678.1) in Romanian sheep affected by lung cancer and showed a high similarity with the UK strain (AF105220.1)., Conclusions: In this study, we confirmed for the first time in Romania the presence of exJSRV in naturally occurring OPA in sheep. Additionally, we described the first report of atypical OPA in Romania, and to the best of our knowledge, in Eastern Europe. Finally, we showed that MGs have a myofibroblastic origin.

The purpose of the present study was to determine the histological and ultrastructural changes in the luminal epithelium of the shell gland associated with natural moulting. Samples of the shell gland from laying (32 weeks old) and moulting (75 weeks old) hens were studied using histological, histochemical and electron microscopic techniques. In addition, TUNEL was used to demonstrate the distribution of apoptotic cells in the luminal epithelium of the shell gland. Autophagy, characterized by the presence of autophagosomes and autolysosomes, was evident in the early stages of degeneration in non-ciliated, ciliated and mitochondrial cells. The intermediate and advanced stages of regression in non-ciliated as well as mitochondrial cells occurred via apoptosis, while both apoptotic and necrotic ciliated cells were observed during the later stages of degeneration. The results of the present study suggest that a synergy of autophagy, apoptosis and necrosis is involved in the involution of the shell gland during natural moulting., (© 2020 Blackwell Verlag GmbH.)

Four cases of a rare melanotic variant of malignant nerve sheath tumour (MNST) in dogs are described. All four cases presented with neurological clinical signs due to multicentric, intradural, intra- and extraparenchymal neoplasms that surrounded the spinal and cranial nerves and infiltrated the adjacent spinal cord and brain. The dogs were young (3 months to 3 years of age), all were female and four different breeds were represented. Characteristic histological features were interweaving fascicles of spindle-shaped cells, sometimes with an architecture reminiscent of Antoni A and B patterns. Some spindle cells showed prominent cytoplasmic melanin pigmentation and such cells were positive by Masson-Fontana stain. Immunohistochemistry performed in three cases was positive for S100 and vimentin, strongly positive for melan A in the melanized cells and negative for glial fibrillary acidic protein and periaxin. Non-melanized cells did not express melan A. Transmission electron microscopy findings in one case were consistent with a peripheral nerve sheath tumour and demonstrated cytoplasmic pre-melanosomes and melanosomes. Melanotic variants of MNSTs are rare in animals with only a solitary report of two previous canine cases in the literature., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

The aim of this research work was to study the histological structure of the pancreatic acini by transmission electron microscope in two avian species, duck and pigeon. The specimens were collected and processed for electron microscopic study. The results showed that the acini of the two avian species were two types; the first one was an electron dense and the second one an electron lucent. The light acinar cells were larger in size than the dark cells. These cells contained centrally located ovoid nuclei with prominent nucleoli and abundant euchromatin. The cytoplasm was electron lucent, with many rough endoplasmic reticulum, polymorphic mitochondria. Numerous zymogen granules were distributed in the basal part and around the nucleus, so these cells considered active cells. The dark acinar cells were characterized by an electron dense cytoplasm. The most prominent cell organelle in these cells were the zymogen granules that appeared in different sizes while other organelles as mitochondria, and rough endoplasmic reticulum were inconspicuous or few, so these cells were considered as inactive cells. The nucleus with indented nuclear membrane located centrally with prominent nucleoli and abundant heterochromatin. Prominent intercellular spaces between the individual acinar cells, as well as well-developed basement membrane separating the electron dense cells and the lumen contained the secretion between acinar cells. It could be concluded that the acinar cells in ducks and pigeons were divided into two types, that is, light and dark acinar cells which mainly attributed to the activity of these cells., (© 2020 Blackwell Verlag GmbH.)

A 26-year and 6-month-old male sika deer that was kept at the Showa Park, Tokyo, Japan, collapsed and died of severe disease wasting and severe tabefaction. Grossly, numerous masses, 0.3-1.0 cm diameter, were dispersed throughout the liver. The multiple masses were composed of tumor cells, which had hypochromatic nuclei and abundant faintly eosinophilic cytoplasm, arranged in nests of various sizes. Immunohistochemically, tumor cells were positive for cytokeratin, chromogranin A, synaptophysin and gastrin. Ultrastructurally, the cytoplasm of the tumor cells contained abundant membrane-bound electron-dense granules. A metastatic lesion was observed in the renal, hepatic and pancreatic lymph nodes. On the basis of these findings, this tumor was diagnosed as a neuroendocrine carcinoma with metastases to the lymph nodes.

A 5-mo-old Bassett Hound-Labrador Retriever cross was autopsied following a bout of lethargy, inappetence, and bleeding gums. Mucous membranes were white, and the small intestine was blue-black; the colon contained black feces. The spleen was swollen, and multiple lymph nodes were enlarged and hemorrhagic. Microscopically, the small intestine had focal crypt cell necrosis and circumferential transmural vasculitis, the latter the cause of infarction and the blue-black coloration. Lymphocytes were necrotic in spleen and lymph nodes, and erythrophagocytosis was present in some nodes. Vasculitis was present in brain, meninges, lung, liver, and kidneys. Electron microscopy revealed aggregates of 15-18 nm round viral particles in damaged crypt cells and in the endothelium of small blood vessels. Electron-dense intracytoplasmic inclusions consisting of paracrystalline-arrayed virus were demonstrated in macrophages in medullary lymph node sinuses. These virions were identified as circovirus, which was confirmed by real-time PCR and sequencing.

Twenty-three fishes were used to study the structure and ultrastructure of interrenal tissue, chromaffin cells and corpuscles of Stannius of Nile tilapia. The interrenal tissue and chromaffin cells are present within the head kidney. The interrenal tissue is arranged in the form of highly convoluted cords, bordered by the lining endothelium of the adjacent sinusoids. It has no connective tissue capsule. The cytoplasm of the interrenal cells contains abundance of mitochondria, vacuoles and smooth endoplasmic reticulum, characterizing of steroid-producing tissues. Two types of chromaffin cells; noradrenaline (NA) cells and adrenaline cells (A) could be recognized by light microscope using chromaffin reaction, as well as by electron microscope they could be distinguished depending on the size and electron density of their granules. The corpuscles of Stannius are two in number and located on the dorsal aspect of the tail kidney. Each corpuscle is surrounded by thick connective tissue capsule. The parenchyma is divided into lobules, each of which is surrounded by distinct basal lamina and has a pseudo lumen. Depending on the presence of secretory granules and the relative abundance of cell organelles, three cell types could be recognized; granular cell, agranular cell (Type I) and agranular cell (Type II). In conclusion, the morphological and ultrastructural analysis of the endocrine tissues of the kidney of Nile tilapia has revealed only one type of interrenal cells, two types of chromaffin cells and three staged-cells of Stannius corpuscles., (© The Author(s) 2019. Published by Oxford University Press on behalf of The Japanese Society of Microscopy. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

An ultrastructural study on the platelets from peripheral blood samples from 20 healthy adult Arabian oryx (Oryx leucocoryx) was performed. Characteristic was the extreme polymorphism of both the platelets, as well as their alpha granules. They vary in size from 100 to 800 nm in diameter and their numbers typically are less than those reported for humans and other animal species. Also, the alpha granules in contrast to those of humans and animals, such as the Arabian tahr, do not have nucleoids. Typically, the oryx platelets exhibit only 1-2 electron-dense bodies per cell and they lack an open canalicular system. Of particular interest is the presence of Type I multivesicular bodies, which have previously only been described in human megakaryocytes and are hypothesized as being intermediate development stages of alpha and dense granules., (© 2019 Blackwell Verlag GmbH.)

Hydropericardium syndrome (HPS) is one of the important emerging diseases causing huge losses to the poultry industry. It affects mainly 3- to 6-week-old broiler chickens and rarely occurs in breeding and laying flocks. Recently, an HPS case was recorded with a sudden heavy mortality in a 100-day-old laying flock. A fowl adenovirus serotype 4 (FAdV-4), named as GDMZ strain, was isolated and identified using polymerase chain reaction coupled with electron microscopy. The animal experiment showed that a mortality of 100% was recorded with hydropericardium as a conspicuous lesion throughout the course of infection. Microscopically, vacuolar changes and intranuclear inclusion bodies were observed in the liver and vacuolar changes were observed in the heart. The complete genome sequence of GDMZ strain was determined to investigate the molecular properties of GDMZ strain. The comparative analysis revealed that the novel Chinese FAdV-4 isolate contained open reading frame (ORF) 19, ORF27, and ORF48 genomic deletions. The phylogenetic analysis revealed that FAdV-4 could be divided into two major clades, of which Chinese FAdV-4 were located at a distinct clade.

Sperm plasma membrane is an essential structure of sperm resistance to freezing. Signs of cryodamage can be visible on the sperm plasma membrane. The aim of our study was to evaluate the appearance of plasma membrane and acrosome in fresh and frozen-thawed chicken sperm using electron and fluorescence microscopy. Semen was collected from 12 sexually mature roosters of Ross PM3 heavy line, diluted with Kobidil + extender with 16% of ethylene glycol (KEG; control) or with KEG in combination with one of following non-permeating cryoprotectants: trehalose (KEG-TRE) or glycine (KEG-GLY). Fluorescence staining was used for detection of the membrane integrity, apoptotic changes and viability (Annexin V, Yo-PRO-1, PI, respectively). Ultrathin sections (70 nm) from samples were prepared to examine sperm head ultrastructure. Freezing process significantly worsened the status of the sperm plasma membranes. In all frozen groups, only about a quarter of the evaluated sperm were graded as class I quality. In the KEG and KEG-GLY groups, about half of sperm had severe plasma membrane damages (III class). In sperm with extensively damaged membranes (III class), the acrosome-sperm head junction was mostly disturbed. The use of trehalose was more beneficial (p < 0.05) for sperm plasma membrane than the use of glycine. In contrast, a decrease (p < 0.05) in the apoptotic sperm ratio (Yo-PRO-1) was noted in the KEG-GLY group when compared to other treatments. In conclusion, we identified different plasma membrane and acrosome damages in cryopreserved chicken sperm. The loss of acrosomes can contribute to diminishing of fertilization ability of cryopreserved chicken sperm., (© 2018 Blackwell Verlag GmbH.)

Macrococcus caseolyticus is generally considered to be a non-pathogenic bacterium that does not cause human or animal diseases. However, recently, a strain of M. caseolyticus (SDLY strain) that causes high mortality rates was isolated from commercial broiler chickens in China. The main pathological changes caused by SDLY included caseous exudation in cranial cavities, inflammatory infiltration, haemorrhages and multifocal necrosis in various organs. The whole genome of the SDLY strain was sequenced and was compared with that of the non-pathogenic JCSC5402 strain of M. caseolyticus. The results showed that the SDLY strain harboured a large quantity of mutations, antibiotic resistance genes and numerous insertions and deletions of virulence genes. In particular, among the inserted genes, there is a cluster of eight connected genes associated with the synthesis of capsular polysaccharide. This cluster encodes a transferase and capsular polysaccharide synthase, promotes the formation of capsules and causes changes in pathogenicity. Electron microscopy revealed a distinct capsule surrounding the SDLY strain. The pathogenicity test showed that the SDLY strain could cause significant clinical symptoms and pathological changes in both SPF chickens and mice. In addition, these clinical symptoms and pathological changes were the same as those observed in field cases. Furthermore, the anti-microbial susceptibility test demonstrated that the SDLY strain exhibits multiple-antibiotic resistance. The emergence of pathogenic M. caseolyticus indicates that more attention should be paid to the effects of this micro-organism on both poultry and public health., (© 2018 Blackwell Verlag GmbH.)

Coccidioidomycosis in nonhuman primates has been sporadically reported in the literature. This study describes 22 cases of coccidioidomycosis in nonhuman primates within an endemic region, and 79 cases of coccidioidomycosis from the veterinary literature are also reviewed. The 22 cases included baboons ( n = 10), macaques ( n = 9), and chimpanzees ( n = 3). The majority died or were euthanized following episodes of dyspnea, lethargy, or neurologic and locomotion abnormalities. The lungs were most frequently involved followed by the vertebral column and abdominal organs. Microscopic examination revealed granulomatous inflammation accompanied by fungal spherules variably undergoing endosporulation. Baboons represented a large number of cases presented here and had a unique presentation with lesions in bone or thoracic organs, but none had both intrathoracic and extrathoracic lesions. Although noted in 3 cases in the literature, cutaneous infections were not observed among the 22 contemporaneous cases. Similarly, subclinical infections were only rarely observed (2 cases). This case series and review of the literature illustrates that coccidioidomycosis in nonhuman primates reflects human disease with a varied spectrum of presentations from localized lesions to disseminated disease.

Background: Blood centrifugation and buffy coats are at the cornerstone of hematology. In mammals, the buffy coat has a layered disposition (from bottom to top) with neutrophils on top of erythrocytes, followed by monocytes/lymphocytes, and platelets. In nonmammals, this distribution is unknown. Recently, the cell tube block (CTB) technique was developed to study the buffy coat, but it was never applied to nonmammal buffy coats., Objectives: This study aimed to evaluate using the CTB technique to study reptilian and avian buffy coats and to propose its use for clinical applications., Methods: Blood from five birds and eight reptiles of different species was obtained to make CTBs that were processed for optical/electron microscopy. H&E, Sirius red, and immunohistochemistry staining against CD3 (to label T lymphocytes) were applied to the CTBs., Results: In birds, the buffy coat had a layered appearance with the granulocyte layer containing granulocytes (heterophils and eosinophils) and nucleated erythrocytes followed by a mononuclear cell layer containing lymphocytes, monocytes, and thrombocytes. In some animals, a nucleated erythrocyte layer was observed admixed with the granulocyte/mononuclear cell layer. A small clot within the buffy coat was seen in seven reptiles, and less definition of layers occurred in reptiles, with only one or two layers. Lymphocytes appeared toward the top of the buffy coat., Conclusions: From a comparative hematology perspective, the buffy coat of mammals differs from that of birds and more from that of reptiles. The CTB technique can be used to study these differences in avian and reptilian hematology, especially to study atypical circulating cells, hemoparasites, or blood cell proportions in health and disease., (© 2018 American Society for Veterinary Clinical Pathology.)

Patellar luxation is abnormal displacement of the patella from the femoral trochlear groove. It is seen primarily in small breed dogs and causes pain and limited mobility of the stifle joint. This study aimed to investigate the relationship among patellar luxation, skin extension, and skin collagen fibril diameter. Nine dogs with patellar luxation and five clinically normal dogs were enrolled in the study. We measured the skin extension and investigated the ultrastructure of the skin and patellofemoral ligament by histopathology and transmission electron microscopy. The mean skin extension in dogs with patellar luxation was 18.5 ± 5.5% which is greater than the reference value (14.5%). Mean skin extension in controls was 8.8 ± 1.7% and was within the normal range. In dogs with patellar luxation, histopathology of the skin and patellofemoral ligament showed sparse and unevenly distributed collagen fibers. Transmission electron microscopy identified poorly organized, irregularly shaped, thin collagen fibrils. Collagen fibril thickness in dogs with patellar luxation was significantly less than fibril thickness in controls (P<0.001). There was a significant negative correlation (ρ= -0.863; P<0.001) between skin collagen fibril diameter and skin extension. Skin extension was correlated with patellar luxation and disease severity. Dogs with patellar luxation, joint dysplasia, and hyperextensible skin appear to be pathologically related. This might represent a phenotype of the Ehlers-Danlos syndrome, a hereditary connective tissue disorder in humans.

Aleutian mink disease virus is the type species in the genus Amdoparvovirus, and in mink and other Mustelidae can cause either subclinical disease or fatal chronic immune stimulation and immune complex disease. The authors describe a novel amdoparvovirus in the endangered red panda ( Ailurus fulgens), discovered using viral metagenomics. The authors analyzed the prevalence, tissue distribution, and disease association by PCR, in situ hybridization, electron microscopy, and histology in a group of 6 red pandas from a single zoological collection. The study incorporates a fecal shedding survey and analysis of tissues from 4 necropsied animals over a 12-year span. The tentatively named red panda amdoparvovirus (RpAPV) was detected in the feces and/or tissues of all animals tested. At necropsy of 1 geriatric animal, infection was associated with pyogranulomatous peritonitis, pancreatitis, and myocarditis. Other animals had detectable low-level viral nucleic acid in lymph nodes and both oral and intestinal epithelium at the time of necropsy. Full-length genome sequences of RpAPV strains from 2 animals had 12% sequence divergence, demonstrating genetic diversity even among in-contact animals. RpAPV is a persistent infection in this cohort of red pandas, and has variable clinical expression.

Renal fibrosis is characterized by glomerulosclerosis and tubulointerstitial fibrosis and its pathogenesis is associated with the activity of mesenchymal cells (fibroblasts), being essentially characterized by a process of excessive accumulation resulting from the deposition of extracellular matrix components. The aim of this study was to characterize the morphological presentation of chronic and fibrotic lesions in the glomerular, tubular, interstitial, and vascular compartments in feline CKD, as well as the possible participation of myofibroblasts in renal fibrotic processes in this species. Cat kidneys were collected and processed according to the conventional techniques for light microscopy, circular polarization, immunohistochemistry, and electron microscopy. Fibrotic alterations were present in all compartments analyzed. The main findings in the glomerular compartment were different degrees of glomerular sclerosis, synechia formation, Bowman's capsule calcification, in addition to glomerular basement membrane thickening and pericapsular fibrosis. The tubulointerstitial compartment had intense tubular degeneration and the immunostaining in tubular cells for mesenchymal cell markers demonstrated the possibility of mesenchymal epithelial transition and consequent involvement of myofibroblasts in the development of interstitial tubule damage. Infiltration of inflammatory cells, added to vessel thickening and fibrosis, demonstrated the severity and role of inflammation in the development and perpetuation of damage. Thus, we may conclude that fibrotic lesions play a relevant role in feline CKD and the mechanism of perpetuation of these lesions need further elucidation regarding the origin and participation of myofibroblasts and consequent mesenchymal epithelial transition in this species., (© 2017 Wiley Periodicals, Inc.)

A missense variant in the autophagy-related ATG4D-gene has been associated with a progressive degenerative neurological disease in Lagotto Romagnolo (LR) dogs. In addition to neural lesions, affected dogs show an extraneural histopathological phenotype characterized by severe cytoplasmic vacuolization, a finding not previously linked with disturbed autophagy in animals. Here we aimed at testing the hypothesis that autophagy is altered in the affected dogs, at reporting the histopathology of extraneural tissues and at excluding lysosomal storage diseases. Basal and starvation-induced autophagy were monitored by Western blotting and immunofluorescence of microtubule associated protein 1A/B light chain3 (LC3) in fibroblasts from 2 affected dogs. The extraneural findings of 9 euthanized LRs and skin biopsies from 4 living affected LRs were examined by light microscopy, electron microscopy, and immunohistochemistry (IHC), using antibodies against autophagosomal membranes (LC3), autophagic cargo (p62), and lysosomal membranes (LAMP2). Biochemical screening of urine and fibroblasts of 2 affected dogs was performed. Under basal conditions, the affected fibroblasts contained significantly more LC3-II and LC3-positive vesicles than did the controls. Morphologically, several cells, including serous secretory epithelium, endothelial cells, pericytes, plasma cells, and macrophages, contained cytoplasmic vacuoles with an ultrastructure resembling enlarged amphisomes, endosomes, or multivesicular bodies. IHC showed strong membranous LAMP2 positivity only in sweat glands. The results show that basal but not induced autophagy is altered in affected fibroblasts. The ultrastructure of affected cells is compatible with altered autophagic and endo-lysosomal vesicular traffic. The findings in this spontaneous disease provide insight into possible tissue-specific roles of basal autophagy.

Background: The European Veterinary Renal Pathology Service (EVRPS) is the first Web-based registry for canine renal biopsy specimens in Europe., Hypothesis/objectives: The aim was to verify whether differences exist between the clinical and laboratory presentation of dogs with nephropathy according to renal pathological findings, as defined by light and electron microscopy of renal biopsy specimens submitted to EVRPS., Animals: Renal biopsy specimens of dogs were collected from the archive of the service (n = 254). Cases were included if both light and electron microscopy were available (n = 162)., Methods: Renal biopsy specimens were classified based on the morphological diagnoses. Thereafter, they were grouped into 3 disease categories, including immune-complex-mediated glomerulonephritis (ICGN), non-immune-complex-mediated GN (non-ICGN), and renal lesions not otherwise specified (RL-NOS). Differences among morphological diagnoses and among disease categories were investigated for clinical and laboratory variables., Results: Serum albumin concentration was lower in dogs with ICGN than in those with non-ICGN (P = 0.006) or RL-NOS (P = 0.000), and the urine protein-to-creatinine ratio (UPC) was significantly higher in ICGN than in the other 2 disease categories. Regarding morphological diagnoses, albumin was significantly lower in amyloidosis (AMY) and membranous (MGN), membranoproliferative (MPGN) or mixed glomerulonephritis (MixGN) than in minimal change disease, primary (FSGS I) or secondary (FSGS II) focal and segmental glomerulosclerosis and juvenile nephropathies (JN). The UPC was higher in MPGN than in FSGS I and FSGS II., Conclusions and Clinical Importance: Dogs with ICGN, in particular MPGN, had higher protein loss than those with non-ICGN or RL-NOS, leading to more severe hypoalbuminemia. Clinical and laboratory differentiation among dogs with the different morphological diagnoses and among dogs with different disease categories was difficult due to overlapping results., (Copyright © 2017 The Authors. Journal of Veterinary Internal Medicine published by Wiley Periodicals, Inc. on behalf of the American College of Veterinary Internal Medicine.)

Swine vesicular disease (SVD) is a highly contagious viral disease that causes vesicular disease in pigs. The importance of the disease is due to its indistinguishable clinical signs from those of foot-and-mouth disease, which prevents international trade of swine and related products. SVD-specific antibody detection via an enzyme-linked immunosorbent assay (ELISA) is the most versatile and commonly used method for SVD surveillance and export certification. Inactivated SVD virus is the commonly used antigen in SVD-related ELISA. A recombinant SVD virus-like particle (VLP) was generated by using a Bac-to-Bac baculovirus expression system. Results of SVD-VLP analyses from electron microscopy, western blotting, immunofluorescent assay, and mass spectrometry showed that the recombinant SVD-VLP morphologically resemble authentic SVD viruses. The SVD-VLP was evaluated as a replacement for inactivated whole SVD virus in competitive and isotype-specific ELISAs for the detection of antibodies against SVD virus. The recombinant SVD-VLP assay produced results similar to those from inactivated whole virus antigen ELISA. The VLP-based ELISA results were comparable to those from the virus neutralization test for antibody detection in pigs experimentally inoculated with SVD virus. Use of the recombinant SVD-VLP is a safe and valuable alternative to using SVD virus antigen in diagnostic assays.

Morphology of important cell organelles (mitochondria, lipid droplets, vacuoles, inclusion bodies and apoptotic bodies) in embryos derived from cows with different body condition score (BCS) was analysed by transmission electron microscopy (TEM). Embryos were recovered on 7th day after the insemination by a standard non-surgical flushing of the uterine horns from superovulated Holstein Friesian cows with BCS 2, 3, 4 and 5. Thereafter, the good quality blastocysts were processed for TEM. The electronograms were evaluated by stereological analysis. The relative volume of lipid droplets in BCS4 and BCS5 embryos increased significantly (18.53 and 22.40%) when compared to BCS3 embryos (5.46%). In the embryos from the BCS4 or BCS5 cows, we observed different morphological patterns of mitochondria, as well as the mitochondria containing vacuoles. BCS4 and BCS5 embryo cell nuclei showed the structure typical for low transcription activity (none or very few reticular nucleoli); also dilated inter-cellular spaces were often observed in these embryos. In conclusion, differences in the ultrastructural morphology of embryos from over-conditioned cows (BCS4 and BCS5), particularly the higher lipid content in the cytoplasm, can be a marker of their low quality, and this fact can be a contributing factor to subfertility in over-conditioned cows., (© 2017 Blackwell Verlag GmbH.)

Neoplasia is usually encountered in the African pygmy hedgehog at a mean age of 3.5 y, and malignancy is common. Myelogenous leukemias are rarely reported in hedgehogs. We describe 3 cases of eosinophilic leukemia in adult, middle-aged (mean age: 2.3 y) hedgehogs, for which prognosis appears grave. In 1 case, attempted treatment was unsuccessful, and in all 3 cases, the disease course was rapid and all died soon after diagnosis. Blood smear evaluation, along with complete blood count, was critical in making the diagnosis in all cases. Luna stain was validated and used to better visualize eosinophils in cytologic and histologic sections. Electron microscopy confirmed the presence of specific granules in hedgehog eosinophils.

Myelination, the ensheathing of neuronal axons by myelin, is important for the proper function of both central and peripheral nervous systems. Various studies have investigated the quantitative parameters of myelination in certain species. Pigs are among the species of which their use as laboratory animals in neuroscience research increased the past few decades. However, there is limited data regarding the myelination process in the pig. Moreover, the maxillary nerve is crucial for Pseudorabies Virus (PrV) neuropathogenesis. In this context, a quantitative analysis of various myelination parameters of the maxillary nerve was performed, during the first 5 weeks of porcine post-natal development, the time period, which exhibits the highest interest for PrV neuropathogenesis. The evaluation was conducted in four groups of uninfected pigs, at the time of birth (group 0w), at the age of 1 week (group 1w), 3 weeks (group 3w) and 5 weeks (group 5w), using toluidine blue staining, immunofluorescence and electron microscopy. Axon and fibre diameter, perimeter and surface, myelin sheath thickness and g-ratio were measured on histological sections transverse to the longitudinal axis of the maxillary nerve. The thickness of myelin sheath was 0.76 μm for group 0w, 0.94 μm for group 1w, 0.98 μm for group 3w and 1.03 μm for group 5w. The g-ratio was 0.529, 0.540, 0.542 and 0.531 for the respective animal groups. The results of this study contribute to the understanding of the myelination process in the pig will be used for the study of PrV effects on the myelination development of newborn piglets' maxillary nerve and may shed new light to their vulnerability to the virus., (© 2016 Blackwell Verlag GmbH.)

Endometritis is a common puerperal disease in livestock. Thus, a practical clinical test for the establishment of diagnosis is needed. We developed a goat model for subclinical endometritis diagnosis by examining cytological, ultrastructural features and molecular expression in cervical discharge collected by cervical cytobrush method. A suspension of E. coli was infused into the uterine horn of goats, and cervical discharge, peripheral blood and endomertrial biopsy samples were collected before inoculation and at 3, 6, 12, 24, 72, 120 and 168 hr post-inoculation (pi). In experimental goats, total leukocytes increased at 6-12 hr pi in hematological examinations, abundant neutrophils were observed in cervical discharge smears, and some micro-villi shortened or were lost from the epithelium at 3 hr pi, with some inflammatory cells infiltrated into the uterine glands and lamina propria. The mRNA expression levels of the Toll-like receptor 4 (TLR4), cytokines and β-defensin-2 increased significantly within 12 hr pi and returned to pre-inoculation levels at 7 day pi. The results showed that subclinical endometritis was simulated in this bacterial infusion goat model, and the evaluation of cervical cytology was in good agreement with molecular changes of cytokines in cervical discharge, hematology and histology. We thus conclude that the simple cervical cytobrush technique can effectively obtain cervical discharge samples for cytology examination in the early diagnosis of subclinical endometritis in goats.

Here we report a pig with amyloid A (AA) amyloidosis associated with Streptococcus suis infection and identification of a unique amyloid sequence in the amyloid deposits in the tissue. Tissues from the 180-day-old underdeveloped pig contained foci of necrosis and suppurative inflammation associated with S. suis infection. Congo red stain, immunohistochemistry, and electron microscopy revealed intense AA deposition in the spleen and renal glomeruli. Mass spectrometric analysis of amyloid material extracted from the spleen showed serum AA 2 (SAA2) peptide as well as a unique peptide sequence previously reported in a pig with AA amyloidosis. The common detection of the unique amyloid sequence in the current and past cases of AA amyloidosis in pigs suggests that this amyloid sequence might play a key role in the development of porcine AA amyloidosis. An in vitro fibrillation assay demonstrated that the unique AA peptide formed typically rigid, long amyloid fibrils (10 nm wide) and the N-terminus peptide of SAA2 formed zigzagged, short fibers (7 nm wide). Moreover, the SAA2 peptide formed long, rigid amyloid fibrils in the presence of sonicated amyloid fibrils formed by the unique AA peptide. These findings indicate that the N-terminus of SAA2 as well as the AA peptide mediate the development of AA amyloidosis in pigs via cross-seeding polymerization.

The New World monkey Aotus spp. (night monkeys) are expected for use of valuable experimental animal with the close species of Saimiri spp. (squirrel monkeys). Saimiri is known to show spontaneous hypercortisolemia, although few reports in Aotus. We compared basic states of blood steroid hormones and histological structure of the adrenal glands in two monkeys. Serum cortisol and ACTH levels were statistically lower in Aotus than Saimiri. Conversely, Aotus adrenocortical area showed significant enlargement, especially at the zona fasciculata. Electron microscopic observation at Aotus fasciculata cells revealed notable accumulation of large lipid droplets and irregular shapes of the mitochondrial cristae. These results suggest potential differences in cellular activities for steroidogenesis between Aotus and Saimiri and experimental usefulness in adrenocortical physiology and pathological models.

Unlabelled: The white sucker Catostomus commersonii is a freshwater teleost often utilized as a resident sentinel. Here, we sequenced the full genome of a hepatitis B-like virus that infects white suckers from the Great Lakes Region of the United States. Dideoxy sequencing confirmed that the white sucker hepatitis B virus (WSHBV) has a circular genome (3,542 bp) with the prototypical codon organization of hepadnaviruses. Electron microscopy demonstrated that complete virions of approximately 40 nm were present in the plasma of infected fish. Compared to avi- and orthohepadnaviruses, sequence conservation of the core, polymerase, and surface proteins was low and ranged from 16 to 27% at the amino acid level. An X protein homologue common to the orthohepadnaviruses was not present. The WSHBV genome included an atypical, presumptively noncoding region absent in previously described hepadnaviruses. Phylogenetic analyses confirmed WSHBV as distinct from previously documented hepadnaviruses. The level of divergence in protein sequences between WSHBV and other hepadnaviruses and the identification of an HBV-like sequence in an African cichlid provide evidence that a novel genus of the family Hepadnaviridae may need to be established that includes these hepatitis B-like viruses in fishes. Viral transcription was observed in 9.5% (16 of 169) of white suckers evaluated. The prevalence of hepatic tumors in these fish was 4.9%, and only 2.4% of fish were positive for both virus and hepatic tumors. These results are not sufficient to draw inferences regarding the association of WSHBV and carcinogenesis in white sucker., Importance: We report the first full-length genome of a hepadnavirus from fishes. Phylogenetic analysis of this genome indicates divergence from genomes of previously described hepadnaviruses from mammalian and avian hosts and supports the creation of a novel genus. The discovery of this novel virus may better our understanding of the evolutionary history of hepatitis B-like viruses of other hosts. In fishes, knowledge of this virus may provide insight regarding possible risk factors associated with hepatic neoplasia in the white sucker. This may also offer another model system for mechanistic research., (Copyright © 2015 Hahn et al.)

Collagen type III glomerulopathy, also known as collagenofibrotic glomerulopathy, is a rare renal disease of unknown pathogenesis. The disease occurs in humans and animals and is characterized by massive glomerular accumulations of collagen type III. In the present study, we describe a Drever dog litter affected by an early onset variant of this glomerular disease, where 4 of 9 puppies developed renal failure within 50 days of age. Necropsy specimens of kidney from the 4 affected cases were studied by light microscopy, electron microscopy, and immunohistochemistry, and characteristic lesions compatible with a diagnosis of collagen type III glomerulopathy were found. In addition, 2 cases showed atypical epithelium in the collecting ducts of the medulla, so-called adenomatoid change. Immunohistochemistry of renal specimens from collagen type III glomerulopathy-affected dogs (n = 10) originating from two different dog strains, the Drever dogs and a mixed-breed strain, demonstrated that the deposited glomerular collagen is composed of a mixture of collagen III and collagen V. The distribution of the collagen V corresponded to the localization of collagen III; however, differences in staining intensity showed that collagen type III is the dominating component. Immunohistochemistry for collagen III (n = 9) and a transmission electron microscopic study (n = 1) showed hepatic perisinusoidal collagen type III deposition in affected cases from both dog strains. This is the first report documenting glomerular accumulations of collagen type V and perisinusoidal liver collagen III deposition in canine collagen type III glomerulopathy., (© The Author(s) 2014.)

A chronic progressive neurological condition in an Alexandrine parrot (Psittacula eupatria) was manifest as intention tremors, incoordination, and seizure activity. Histology revealed large eosinophilic bodies throughout the central nervous system, and electron microscopy demonstrated that these bodies were greatly expanded axons distended by short filamentous structures that aggregated to form long strands. The presence of periodic acid-Schiff-positive material within the neuronal bodies of Purkinje cells and ganglionic neurons is another distinctive feature of this disease. The histological features of this case display some features consistent with giant axonal neuropathy as reported in humans and dogs. Based on investigation of the lineage in this case, an underlying inherited defect is suspected, but some additional factor appears to have altered the specific disease presentation in this bird., (© 2015 The Author(s).)

An avian paramyxovirus (APMV) isolated from goose feces (APMV/Shimane67) was biologically, serologically and genetically characterized. APMV/Shimane67 showed typical paramyxovirus morphology on electron microscopy. On hemagglutination inhibition test, antiserum against APMV/Shimane67 revealed low reactivity with other APMV serotypes and vice versa. The fusion (F) protein gene of APMV/Shimane67 contained 1,638 nucleotides in a single open reading frame encoding a protein of 545 amino acids. The cleavage site of F protein contained a pair of single basic amino acid (VRENR/L). The nucleotide and deduced amino acid sequences of the F gene of APMV/Shimane67 had relatively low identities (42.9-62.7% and 28.9-67.3%, respectively) with those of other APMVs. Phylogenetic analysis showed that APMV/Shimane67 was related to NDV, APMV-9 and APMV-12, but was distinct from those APMV serotypes. These results suggest that APMV/Shimane67 is a new APMV serotype, APMV-13.

In this study, we investigated recent sheep pox outbreaks that occurred in Ononsky and Borzunsky regions of Zabajkalskij kray of Russia. The outbreaks involved in 2756 animals of which 112 were infected and 3 were slaughtered. Samples of injured skin of infected sheep were analysed by electron microscopy and CaPV-specific P32 gene amplification. Following sequence analysis of entire P32 gene showed that both specimens were identical to the sequence of several sheep poxvirus isolates from China and India. The close location of China to the last decade's Russian outbreaks suggest that possible future outbreaks in Russia could occur along the border regions with countries where sheep and goat pox are not controlled., (© 2013 Blackwell Verlag GmbH.)