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Extracellular vesicles (EVs) play a crucial role in cell-to-cell communication and have great potential as efficient delivery vectors. However, a better understanding of EV in vivo behavior is hampered by the limitations of current imaging tools. In addition, chemical labels present the risk of altering the EV membrane features and, thus, in vivo behavior. 19F-MRI is a safe bioimaging technique providing selective images of exogenous probes. Here, we present the first example of fluorinated EVs containing PERFECTA, a branched molecule with 36 magnetically equivalent 19F atoms. A PERFECTA emulsion is given to the cells, and PERFECTA-containing EVs are naturally produced. PERFECTA-EVs maintain the physicochemical features, morphology, and biological fingerprint as native EVs but exhibit an intense 19F-NMR signal and excellent 19F relaxation times. In vivo 19F-MRI and tumor-targeting capabilities of stem cell-derived PERFECTA-EVs are also proved. We propose PERFECTA-EVs as promising biohybrids for imaging biodistribution and delivery of EVs throughout the body.

In collaboration with the American College of Veterinary Pathologists

Five adult Saanen goats received a single oral dose of Heterophyllaea pustulata containing 42.25 μg/kg rubiadin (anthraquinone) and 3 adult goats were untreated controls. All goats were exposed to sunlight and sequential ear skin biopsies were collected before treatment and at 32 hours, 3 days, 8 days, and 15 days after treatment. Changes at 32 hours after dosing included epidermal spongiosis, single cell death and acantholysis, an increased BAX/BCL-2 protein ratio, and dermal edema. Lesions at day 3 included epidermal and adnexal necrosis, crust formation, and acanthosis. Acanthosis, hyperkeratosis, and dermal fibrosis and neovascularization were present at day 15. The pro-apoptotic (BAX)/anti-apoptotic (BCL-2) protein ratio increased at 32 hours, whereas epidermal and dermal PCNA immunolabeling increased between days 8 and 15 after treatment. The cutaneous lesions were consistent with sunlight-induced damage, and the occurrence in treated but not control goats indicates photosensitization.

Lesions caused by trombiculid mite larvae ('chiggers') in birds have been inadequately described. A juvenile red-legged partridge (Alectoris rufa) presented with multifocal, crater-like lesions of necrotizing dermatitis, which often contained multiple orange mites. Microscopically, there was a nodular necrotizing and pyogranulomatous dermatitis with occasional intralesional arthropods. Histologically, the mites were readily identified by the presence of stylostomes within the necrotic areas. Morphological and morphometrical studies on isolated mites identified them as Neoschoengastia (Hypogastia) simonovichi. This is the first report of N. simonovichi as an aetiological agent of severe trombiculosis in wild birds in Western Europe. Infestation with this parasite requires further study to understand its role in animal and human trombiculosis and its potential role as a vector of infectious, including zoonotic, agents.

Pulmonary affection is by far the most severe and widespread disease form caused by Small Ruminant Lentiviruses (SRLV) in sheep. The infection implies direct contact between animals and the most important infection route is by the respiratory tract. Disease is insidious and might be seen in one-year-old animals onwards. Non-productive dyspnea is the most common clinical sign and interstitial pneumonia with increased lung size and volume are the pathologic landmarks. Early detection of infected animals is a key step for the diagnosis and it is best achieved by third generation ELISAs that include a multistrain design to detect divergent genotypes. Differential diagnosis must be established with ovine pulmonary adenocarcinoma, the visceral form of ovine caseous lymphadenitis, ovine respiratory complex, gangrenous pneumonia and bacterial and parasitic (Dyctiocaulus filaria, Oestrus ovis) infections. Control is achieved by combining serological and management measures and is based on total or selective culling of infected animals and replacement with uninfected lambs. Serological segregation and replacement with the offspring of seronegative ewes is also a suitable option for control. Selection of resistant sheep might be a future option. Pulmonary affection by SRLV in sheep must be taken into account in the face of a chronic, non-productive dyspnea.

Background: Long non-coding RNAs (lncRNAs) are involved in several immune processes, including the immune response to vaccination, but most of them remain uncharacterised in livestock species. The mechanism of action of aluminium adjuvants as vaccine components is neither not fully understood. Results: We built a transcriptome from sheep PBMCs RNA-seq data in order to identify unannotated lncRNAs and analysed their expression patterns along protein coding genes. We found 2284 novel lncRNAs and assessed their conservation in terms of sequence and synteny. Differential expression analysis performed between animals inoculated with commercial vaccines or aluminium adjuvant alone and the co-expression analysis revealed lncRNAs related to the immune response to vaccines and adjuvants. A group of co-expressed genes enriched in cytokine signalling and production highlighted the differences between different treatments. A number of differentially expressed lncRNAs were correlated with a divergently located protein-coding gene, such as the OSM cytokine. Other lncRNAs were predicted to act as sponges of miRNAs involved in immune response regulation. Conclusions: This work enlarges the lncRNA catalogue in sheep and puts an accent on their involvement in the immune response to repetitive vaccination, providing a basis for further characterisation of the non-coding sheep transcriptome within different immune cells., This research was supported by the Spanish Ministry of Economy and Industry project number AGL2013-49137-C3 to BMJ and University of the Basque Country (UPV/EHU) predoctoral fellowships to MB-A (PIF17/ 306) and EV-M ( PIF15/361) and a postdoctoral fellowship to NA (ESP-COC16/43).

Five adult Saanen goats received a single oral dose of

The use of vaccines has proven to be very effective in controlling and eradicating infectious diseases, both in veterinary and human medicine; however, vaccines can be also the source of an array of problems caused by procedures such as overimmunization. Bluetongue, an orbiviral disease that affects ruminants, is best controlled by the use of inactivated vaccines. During the last years of the past decade, these vaccines were applied all over Europe to control the spreading of the disease, a goal that was accomplished; however, at the same time, several adverse effects related to the vaccination were reported. Especially in sheep, this vaccination campaign brought out a new cachectic and neurologic disease with harmful consequences for the ovine industry. This disease is now recognized as the ovine version of the autoimmune/inflammatory syndrome induced by adjuvants (ASIA syndrome) and poses an immense challenge in veterinary medicine, immunology, and vaccinology.

The use of vaccines including aluminum (Al)–based adjuvants is widespread among small ruminants and other animals. They are associated with the appearance of transient injection site nodules corresponding to granulomas. This study aims to characterize the morphology of these granulomas, to understand the role of the Al adjuvant in their genesis, and to establish the presence of the metal in regional lymph nodes. A total of 84 male neutered lambs were selected and divided into 3 treatment groups of 28 animals each: (1) vaccine (containing Al-based adjuvant), (2) adjuvant-only, and (3) control. A total of 19 subcutaneous injections were performed in a time frame of 15 months. Granulomas and regional lymph nodes were evaluated by clinicopathological means. All of the vaccine and 92.3% of the adjuvant-only lambs presented injection-site granulomas; the granulomas were more numerous in the group administered the vaccine. Bacterial culture in granulomas was always negative. Histologically, granulomas in the vaccine group presented a higher degree of severity. Al was specifically identified by lumogallion staining in granulomas and lymph nodes. Al median content was significantly higher (P < .001) in the lymph nodes of the vaccine group (82.65 μg/g) compared with both adjuvant-only (2.53 μg/g) and control groups (0.96 μg/g). Scanning transmission electron microscopy demonstrated aggregates of Al within macrophages in vaccine and adjuvant-only groups. In these two groups, Al-based adjuvants induce persistent, sterile, subcutaneous granulomas with macrophage-driven translocation of Al to regional lymph nodes. Local translocation of Al may induce further accumulation in distant tissues and be related to the appearance of systemic signs.

Aluminum (Al) hydroxide is an effective adjuvant used in sheep vaccines. However, Al-adjuvants have been implicated as potential contributors to a severe wasting syndrome in sheep&mdash, the so-called ovine autoimmune-inflammatory syndrome induced by adjuvants (ASIA syndrome). This work aimed to characterize the effects of the repetitive injection of Al-hydroxide containing products in lambs. Four flocks (Flocks 1&ndash, 4, n = 21 each) kept under different conditions were studied. Three groups of seven lambs (Vaccine, Adjuvant-only, and Control) were established in each flock. Mild differences in average daily gain and fattening index were observed, indicating a reduced growth performance in Vaccine groups, likely related to short-term episodes of pyrexia and decreased daily intake. Clinical and hematological parameters remained within normal limits. Histology showed no significant differences between groups, although there was a tendency to present a higher frequency of hyperchromatic, shrunken neurons in the lumbar spinal cord in the Adjuvant-only group. Although Al-hydroxide was linked to granulomas at the injection site and behavioral changes in sheep, the results of the present experimental work indicate that injected Al-hydroxide is not enough to fully reproduce the wasting presentation of the ASIA syndrome. Other factors such as sex, breed, age, production system, diet or climate conditions could play a role.

Small Ruminant Lentiviruses (SRLV) are highly prevalent retroviruses with significant genetic diversity and antigenic heterogeneity that cause a progressive wasting disease of sheep called Maedi-visna. This work provides a systematic review and meta-analysis of the last 40 years (1981–2020) of scientific publications on SRLV individual and flock prevalence. Fifty-eight publications and 314 studies were included. Most articles used a single diagnostic test to estimate prevalence (77.6%), whereas articles using three or more tests were scarce (6.9%). Serological tests are more frequently used than direct methods and ELISA has progressively replaced AGID over the last decades. SRLV infection in sheep is widespread across the world, with Europe showing the highest individual prevalence (40.9%) and being the geographical area in which most studies have been performed. Africa, Asia, and North America show values between 16.7% to 21.8% at the individual level. South and Central America show the lowest individual SRLV prevalence (1.7%). There was a strong positive correlation between individual and flock prevalence (ρ = 0.728, p ≤ 0.001). Despite the global importance of small ruminants, the coverage of knowledge on SRLV prevalence is patchy and inconsistent. There is a lack of a gold standard method and a defined sampling strategy among countries and continents.

Bovine viral diarrhoea virus (BVDV) is a pestivirus that affects both cattle and sheep, causing an array of clinical signs, which include abortions and malformations in the offspring. Manufacturing of modified live virus (MLV) vaccines often includes the use of bovine-derived products, which implies a risk of contamination with viable BVDV. Recently, the circulation of a specific strain of BVDV 2b among Spanish sheep flocks, associated with outbreaks of abortions and malformations, and whose origin was not determined, has been observed. On February 2018, a MLV orf vaccine was applied to a 1, 600 highly prolific sheep flock in the Northeast of Spain that included 550 pregnant ewes. In May 2018, during the lambing season, an unusual high rate (72.7%) of abortions, stillbirths, congenital malformations and neurological signs in the offspring was observed. It was estimated that about 1, 000 lambs were lost. Three 1- to 3-day-old affected lambs and a sealed vial of the applied vaccine were studied. Lambs showed variable degrees of central nervous system malformations and presence of pestiviral antigen in the brain. Molecular studies demonstrated the presence of exactly the same BVDV 2b in the tissues of the three lambs and in the orf vaccine, thus pointing to a pestivirus contamination in the applied vaccine as the cause of the outbreak. Interestingly, sequencing at the 5'-untranslated region-(UTR) of the contaminating virus showed a complete match with the virus described in the previously reported outbreaks in Spain, thus indicating that the same contaminated vaccine could have also played a role in those cases. This communication provides a clear example of the effects of the application of this contaminated product in a sheep flock. The information presented here can be of interest in putative future cases of suspected circulation of this or other BVDV strains in ruminants.

Aluminum (Al)-based salts are widely used adjuvants in ruminants and other species to strengthen the immune response elicited against vaccine antigen(s). However, they can lead to the formation of long-lasting granulomas composed of abundant activated macrophages. Small ruminant lentiviruses (SRLV) are widely distributed macrophage-tropic retroviruses that cause persistent infections in sheep and goats. Infected monocytes/macrophages and dendritic cells establish an inflammatory microenvironment that eventually leads to clinical manifestations. The aim of this work was to study the effect of Al-induced granulomas in the replication and pathogenesis of SRLV. Eleven adult, naturally SRLV-infected sheep showing clinical arthritis were distributed in vaccine (n = 6), adjuvant-only (n = 3), and control (n = 2) groups and inoculated with commercial Al-based vaccines, Al hydroxide adjuvant alone, or phosphate-buffered saline, respectively. In vitro studies demonstrated viral replication in Al-induced granulomas in 5 out of 10 sheep. Immunohistochemistry (IHC) evinced granular, intracytoplasmic SRLV presence in macrophages within granulomas. Viral sequences obtained from granulomas, blood monocytes, and other tissues were highly similar in most animals, suggesting virus circulation among body compartments. However, notable differences between isolated strains in granulomas and other tissues in specific animals were also noted. Interestingly, the B2 subtype was the most commonly found SRLV genotype, reaching a wider body distribution than previously described. Recombination events between genotypes B2 and A3 along the gag region were identified in two sheep. Our results indicate that Al-hydroxide-derived granulomas may represent an ideal compartment for SRLV replication, perhaps altering natural SRLV infection by providing a new, suitable target tissue. IMPORTANCE Granulomas are inflammation-derived structures elicited by foreign bodies or certain infections. Aluminum adjuvants included in vaccines induce granulomas in many species. In sheep, these are persistent and consist of activated macrophages. Small ruminant lentiviruses (SRLV), which are macrophage-tropic lentiviruses, cause a chronic wasting disease affecting animal welfare and production. Here, we studied the occurrence of SRLV in postvaccination granulomas retrieved from naturally infected ewes after vaccination or inoculation with aluminum only. SRLV infection was confirmed in granulomas by identification of viral proteins, genomic fragments, and enzymatic activity. The infecting SRLV strain, previously found exclusively in carpal joints, reached the central nervous system, suggesting that occurrence of SRLV in postvaccination granulomas may broaden tissue tropism. SRLV recombination was detected in inoculated animals, a rare event in sheep lentiviruses. Potentially, virus-host interactions within granulomas may modify viral pathogenesis and lead to more widespread infection., This work was funded by grants from the Spanish Ministry of Economy, Industry and Competitiveness (AGL2013-49137-C3-1-R and AGL2013-49137-C3-2-R), the Ministry of Science, Innovation and Universities (RTI2018-096172-B-C31 and RTI2018-096172-B-C33), and the Recognized Research Groups of Government of Aragón (A17_17R, Animal Health and Reproduction). I.E. was a PhD student funded by the Universidad Pública de Navarra. R.d.M. was a PhD student funded by the Department of Innovation, Research and University of Aragón. J.A. and A.R.-L. were PhD students funded by the Spanish Ministry of Science, Innovation and Universities (formerly Spanish Ministry of Education).

Supplemental Material, sj-pdf-1-vet-10.1177_0300985821999322 for Photosensitization Induced by Heterophyllaea pustulata in Goats: Sequential Development of Skin Lesions by Juan F. Micheloud, Lluís Luján, Luis A. Colque-Caro, Susana C. Núñez-Montoya, Claudio G. Barbeito, Enrique L. Portiansky and Eduardo J. Gimeno in Veterinary Pathology

O artigo objetiva revisar alguns dos distúrbios do trato respiratório inferior de maior ocorrência em pequenos ruminantes. Sua rele- vância no manejo sanitário está relacionada à queda no desempenho produtivo, mortalidade, redução do ganho de peso e qualidade da carcaça de cordeiros/cabritos produzidos, e aumento da taxa de descarte e reposição. Aqui, apresentamos a pneumonia por aspiração, complexo respiratório ovino (CRO), adenocarcinoma pulmonar ovino (APO), lentiviroses de pequenos ruminantes (LVPR), pneu- monia parasitária e fúngica, assim como as formas pulmonares da linfadenite caseosa (LC) e das micoplasmoses. A pneumonia por aspiração é uma infecção comumente causada pela inalação de materiais estranhos, produzindo inflamação e necrose do parênquima pulmonar. O CRO é um processo que envolve uma variedade de interações hospedeiro-patógeno-ambiente, em que os mecanismos imunológicos e fisiológicos do hospedeiro interagem com vários agentes etiológicos, incluindo vírus, bactérias, além de fatores ambientais ou estressores. A forma pulmonar da LC comumente causa lesões nos linfonodos mediastinais e no parênquima pulmonar, produzindo sinais clínicos respiratórios, principalmente a tosse responsável pela transmissão da doença por aerossol. O APO é um câncer de pulmão, contagioso, causado pelo retrovírus jaagsiekte ovino, que induz a transformação de células epiteliais secretoras do trato respiratório distal. A afecção pulmonar pela LVPR é a forma de doença mais grave na espécie ovina. A infecção ocorre pelo contato direto entre animais e a via de infecção mais importante é pelo trato respiratório. A pneumonia parasitária e fúngica são menos frequentes, mas relevantes dentro do diagnóstico diferencial das enfermidades respiratórias. Dada a natureza multifacetada dessas doenças, as práticas de manejo (por exemplo, melhorias nas habitações, implementação de medidas de biossegurança e abate seletivo de animais infectados) precisam ser consideradas para sua prevenção; como as condições ambientais também desempenham um papel no desenvolvimento dessas doenças, elas devem ser levadas em consideração. Por fim, outras medidas sanitárias adequadas como protocolos vacinais contra alguns agentes visando a melhora na resposta imune animal também ajudam no controle destas doenças. e article aims to review some of the lower respiratory tract disorders of major occurrence in small ruminants. eir relevance within the health management are related to the drop in productive performance, reduction in weight gain and quality of the carcass of lambs/kids produced, and increase in the mortality, culling and replacement rate. Herein we examine the aspiration pneumonia, ovine respiratory complex (ORC), ovine pulmonary adenocarcinoma (OPA), small ruminant lentiviruses (SRLV ), parasitic and fungal pneumonia, as well as the pulmonary form of caseous lymphadenitis (CLA) and mycoplasmosis. Aspiration pneumonia is an infection commonly caused by inhalation of foreign materials, producing inflammation and necrosis of the lung parenchyma.e ORC is a process involving a range of host-pathogen-environment interactions,where host immunological and physiological mechanisms interact with multiple etiological agents including viruses, bacteria, plus environmental factors or stressors. Pulmonary form of CLA commonly causes lesions in mediastinal lymph nodes and lung parenchyma, producing respiratory clinical signs, particularly the coughing that is responsible for transmission of the disease by aerosol. OPA is a contagious lung cancer of sheep caused by jaagsiekte sheep retrovirus, which induces the transformation of secretory epithelial cells of the distal respiratory tract. Pulmonary affection due to SRLV is the most severe disease form in sheep species. e infection occurs by direct contact between animals and the most important infection routes are lactogenic and respiratory. Parasitic and fungal pneumonia are less frequent, but relevant in the differential diagnosis of respiratory diseases. Given the multi-faceted nature of these diseases, management practices (e.g., housing improvements, implementation of biosecurity provisions and selective culling of infected animals) need to be considered for their prevention; as environmental conditions also play a role in development of these diseases, they should be taken into account. Lastly, other appropriate health measures, such as vaccine protocols against some agents aiming at improving the animal immune response, also help in the control of these diseases.

Aluminium (Al) hydroxide use as adjuvant induces local formation of long‐lasting subcutaneous granulomas in sheep. Macrophages within these granulomas have been identified as a new small ruminant lentivirus (SRLV) replication site in naturally infected animals. Diagnosis of Al hydroxide‐induced granulomas in sheep is mostly based on postmortem observations but little information is available on in vivo detection. Computed tomography (CT) is used for studying these reactions in other animal species. To determine if CT could be a tool for in vivo diagnosis and research of subcutaneous Al hydroxide‐induced granulomas in sheep. A retrospective survey on thoracic CT scans was performed on 46 adult sheep. Analysis included absence or presence, number and location of subcutaneous nodules. Thoracic CT scans and pathological studies were prescribed to two further sheep. Single or multiple subcutaneous nodules were detected in 26 (56.52%) sheep. One or two nodules per animal were most often observed (36.95%). Size ranged between 1.5 and 4.5 cm. Pre‐contrast two‐dimensional (2D) CT images showed focal or multifocal increases in subcutaneous tissue thickness. Post‐contrast 2D CT images revealed hypointense areas in the centre. Histopathology indicated the presence of granulomas composed by a large number of activated macrophages, surrounding a central core of necrosis. Large intracytoplasmic Al‐positive aggregates were demonstrated by lumogallion staining. CT is a useful tool to detect subcutaneous Al hydroxide‐induced granulomas in vivo in sheep. CT provides a diagnostic and research tool that can be very useful in future works in Al hydroxide‐induced pathology, SRLV infection, or both., We describe the usefulness of CT for in vivo diagnose of subcutaneous granulomas in sheep. CT is an excellent tool to advance the diagnosis of subcutaneous Al‐induced granulomas in vivo and to boost further research in SRLV infections in sheep.

Small ruminant lentiviruses (SRLVs) are endemic in most areas of Europe, causing a chronic infection and a multisystemic disease affecting the udder, carpal joints, lungs, and central nervous system. Due to the lack of treatments and protective vaccination strategies, infection control is focused on the identification of infected animals through serological or molecular techniques. However, antigenic and genetic heterogeneity of SRLVs represent a clear drawback for diagnosis. Infected animals may present lower animal production parameters such as birth weight or milk production and quality, depending on productive systems considered and, likely, to the diagnostic method applied. In this study, four sheep flocks dedicated to dairy or meat production were evaluated using three different ELISA and two PCR strategies to classify animal population according to SRLV infection status. Productive parameters were recorded along one whole lactation or reproductive period and compared between positive and negative animals. SRLV was present in 19% of the total population, being unequally distributed in the different flocks. Less than half of the infected animals were detected by a single diagnostic method, highlighting the importance of combining different diagnostic techniques. Statistical analysis employing animal classification using all the diagnostic methods associated lambing size, lamb weight at birth, and daily weight gain with SRLV infection status in meat flocks. Milk production, somatic cell count, fat, and protein content in the milk were associated with SRLV infection in dairy flocks, to a greater extent in the flock showing higher seroprevalence. A multi-platform SRLV diagnostic strategy was useful for ensuring correct animal classification, thus validating downstream studies investigating production traits., This research was funded by the Spanish Ministry of Science, Innovation and Universities, grant number RTI2018-096172-BC31, and by the Government of Navarra and European Union ERDF program, grant number PDR (210160006). LD and IE were funded by Universidad Pública de Navarra. RR was supported by the Spanish Ministry of Science and Innovation Ramón y Cajal contract. Authors acknowledge APC funding by the CSIC Open Access Publication Support Initiative through its Unit of Information Resources for Research (URICI). RD was a Ph.D. student funded by the Department of Innovation, Research and University of Aragón.

The use of vaccines containing aluminum (Al) adjuvants is widespread in ovine production. Al adjuvants induce an effective immune-response but lead to the formation of post-vaccination granulomas from which Al can disseminate. This work aims to study the accumulation of Al in the central nervous system of sheep subcutaneously inoculated with Al-hydroxide containing products. Lumbar spinal cord and parietal lobe from 21 animals inoculated with 19 doses of Vaccine (n = 7), Adjuvant-only (n = 7) or phosphate-buffered saline as Control (n = 7) were analyzed with transversely heated graphite furnace atomic absorption spectroscopy and lumogallion staining for Al analytical measurements and Al tisular localization respectively. In the lumbar spinal cord, Al median content was higher in both the Adjuvant-only and Vaccine group (p = .001) compared with the Control group. Animals of the Adjuvant-only group showed the higher individual measurements in the lumbar spinal cord (14.36 μg/g and 7.83 μg/g). In the parietal lobe, Al median content tended to be higher in the Adjuvant-only group compared with Control group (p = .074). Except for three replicates of the Adjuvant-only group, Al content was always below 1 μg/g. In the lumbar spinal cord, lumogallion-reactive Al deposits were more abundant in the gray matter than in the white matter in both Vaccine (p = .034) and Adjuvant-only groups (p = .017) and Al deposits were mostly associated with glial-like cells (p = .042). In the parietal lobe, few Al deposits, which were sometimes related to blood vessels, were found. In sheep, Al-hydroxide adjuvants inoculated in the subcutaneous tissue selectively accumulate in the lumbar spinal cord., RM is a PhD student funded by the Department of Innovation, Research and University of Aragon, Spain. JA and ARL are PhD students funded by the Spanish Ministry of Science, Innovation and Universities (formerly Spanish Ministry of Education). This work was funded by grants from the Spanish Ministry of Economy, Industry and Competitiveness (AGL2013-49137-C3-1-R and RTI2018-096172-B-C33), the Ministry of Science, Innovation and Universities (RTI2018-096172-B-C31 and RTI2018-096172-B-C33) and the Government of Aragón (A17_17R, Animal Health and Reproduction).

Aluminium hydroxide adjuvants are crucial for livestock and human vaccines. Few studies have analysed their effect on the central nervous system in vivo. In this work, lambs received three different treatments of parallel subcutaneous inoculations during 16 months with aluminium-containing commercial vaccines, an equivalent dose of aluminium hydroxide or mock injections. Brain samples were sequenced by RNA-seq and miRNA-seq for the expression analysis of mRNAs, long non-coding RNAs and microRNAs and three expression comparisons were made. Although few differentially expressed genes were identified, some dysregulated genes by aluminium hydroxide alone were linked to neurological functions, the lncRNA TUNA among them, or were enriched in mitochondrial energy metabolism related functions. In the same way, the miRNA expression was mainly disrupted by the adjuvant alone treatment. Some differentially expressed miRNAs had been previously linked to neurological diseases, oxidative stress and apoptosis. In brief, in this study aluminium hydroxide alone altered the transcriptome of the encephalon to a higher degree than commercial vaccines that present a milder effect. The expression changes in the animals inoculated with aluminium hydroxide suggest mitochondrial disfunction. Further research is needed to elucidate to which extent these changes could have pathological consequences., This work was supported by the Spanish Ministry of Economy grant [MINECO project AGL2013-49137-C3 to BMJ, LL and DA]; University of the Basque Country (UPV/EHU) predoctoral fellowships [PIF15/361 to EV-M and PIF17/306 to MB-A]; and University of the Basque Country (UPV/EHU) postdoctoral fellowship [ESPDOC16/43 to NA].

Wound healing is a complex and costly public health problem that should be timely addressed to achieve a rapid and adequate tissue repair avoiding or even eliminating potential pathogenic infection. Chronic infected non-healing wounds represent a serious concern for health care systems. Efficient wound dressings with tailored therapy having the best response and highest safety margin for the management of chronic non-healing wounds are still needed. The use of novel wound dressing materials has emerged as a promising tool to fulfil these requirements. In this work, asymmetric electrospun polycaprolactone (PCL)-based nanofibers (NFs) were decorated with electrosprayed poly(lactic-co-glycolic acid) microparticles (PLGA MPs) containing the natural antibacterial compound thymol (THY) in order to obtain drug eluting antimicrobial dressings having sustained release. The synthesized dressings successfully inhibited the in vitro growth of Staphylococcus aureus ATCC 25923, showing also at the same doses cytocompatibility on human dermal fibroblasts and keratinocyte cultures after treatment for 24 h, which was not observed when using free thymol. An in vivo murine excisional wound splinting model, followed by the experimental infection of the wounds with S. aureus and their treatment with the synthesized dressings, pointed to the reduction of the bacterial load in wounds after 7 days, though the total elimination of the infection was not reached. The findings indicated the relevance of the direct contact between the dressings and the bacteria, highlighting the need to tune their design considering the wound surface and the nature of the antimicrobial cargo contained.

Sheep health management strategies often include the use of aluminum (Al)-containing vaccines. These products were associated with the appearance of the ovine autoimmune/inflammatory syndrome induced by adjuvants (ASIA syndrome), which included an array of ethological changes in the affected animals. The aim of this pilot study was to investigate cognitive and behavioral changes in sheep subjected to a protocol of repetitive inoculation with Al-containing products. Twenty-one lambs were assigned to three groups (n = 7 each): Control, Adjuvant-only, and Vaccine. Vaccine group was inoculated with commercial Al- hydroxide containing vaccines; Adjuvant-only group received the equivalent dose of Al only (Alhydrogel®), and Control group received Phosphate-buffered saline. Sixteen inoculations were administered within a 349-day period. Ethological changes were studied in late summer (7 inoculations) and mid-winter (16 inoculations). Animals in Vaccine and Adjuvant-only groups exhibited individual and social behavioral changes. Affiliative interactions were significantly reduced, and aggressive interactions and stereotypies increased significantly. They also exhibited a significant increase in excitatory behavior and compulsive eating. There were increased levels of stress biomarkers in these two groups. In general, changes were more pronounced in the Vaccine group than they were in the Adjuvant-only group. Some changes were already significant in summer, after seven inoculations only. This study is the first to describe behavioral changes in sheep after having received repetitive injections of Al-containing products, and may explain some of the clinical signs observed in ovine ASIA syndrome., J. Asín was a PhD student funded by the Ministry of Education, Culture and Sports (MECD). R. Reina was supported by a “Ramón y Cajal” contract from the Spanish Ministry of Economy, Industry and Competitiveness. This work was funded by several grants from the Spanish Ministry of Economy, Industry and Competitiveness (AGL2013-49137-C3-1-R and AGL2013-49137-C3-2-R) and the Ministry of Science, Innovation and Universities (RTI2018-096172-B-C31 and RTI2018-096172-B-C33).

Summary A 20-year-old male brown bear (Ursus arctos) with a 20 × 25 cm necrotic mass adjacent to the trachea was diagnosed as having an anaplastic thyroid carcinoma. Metastases were observed in the lungs and one adrenal gland and, histologically, these had anaplastic and follicular carcinoma patterns, respectively. E-cadherin labelling was observed in the adrenal mass only, while N-cadherin immunolabelling was detected in the thyroid gland and lung masses. Thyroid-specific markers (thyroid transcription factor-1, thyroglobulin) were expressed in the adrenal gland metastasis. This case illustrates an example of a primary epithelial–mesenchymal transition (EMT) enabling metastasis to distant organ sites, followed by a mesenchymal–epithelial transition within the adrenal gland microenvironment, allowing invasion and reacquisition of thyroid epithelial cell features. EMTs help to understand the phenomenon of carcinoma cell plasticity in enabling colonization and growth of metastases.

Trabajo presentado en el Joint Congress of the Veterinary Pathology and Veterinary Clinical Pathology, celebrado en Arnhem (Países Bajos), del 25 al 28 de septiembre de 2019, Ovine management systems usually involve prophylactic vaccination. Most ovine vaccines use aluminium hydroxide as adjuvant. In sheep, aluminium hydroxide promotes an effective immune response, but also induces local granulomatous reactions at the injection site. Aluminium can reach the regional lymph node. The aim of this work was to describe the translocation of subcutaneously inoculated aluminium-based adjuvants to the central nervous system (CNS).

Infection of orthopedic devices is a major complication in the postsurgical period generating important health issues and economic consequences. Prevention strategies could be based on local release of antibiotics from the orthopedic device itself to avoid adhesion and growth of bacteria. The purpose of this work is to demonstrate the efficiency to prevent these infections by a cefazolin-eluting, perforated stainless steel implant in an in vivo ovine model. The device was placed in the tibia of sheep, one group receiving cefazolin-loaded implants whereas the control group received empty implants. All implants were experimentally infected by direct inoculation of Staphylococcus aureus ATCC 6538. In vitro cytotoxicological studies were also performed to check the effect of antibiotic on cell viability, integrity, and cycle. Results showed that sheep receiving cefazolin-loaded devices were able to avoid implant-associated infections, with normal tissue healing process. The antibiotic release followed a local concentric pattern as demonstrated by high-performance liquid chromatography detection in tissues. The in vitro results indicate the lack of relevant cytotoxic effects for the maximum antibiotic concentration released by the device. These results demonstrate the efficiency and safety of cefazolin-eluting implants in an ovine model to prevent early postsurgical infections of orthopedic devices. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 106B: 1976-1986, 2018.

et al., The main challenge of cancer treatment is to avoid or minimize systemic side effects in off-target tissues. Mesenchymal stem cells (MSCs) can be used as therapeutical carriers because of their ability to migrate and incorporate into inflammation areas including tumors. Here, this homing ability is exploited by carrying therapeutic nanoparticles (Hollow Gold Nanoparticles (HGNs)) following a >Trojan-horse> strategy. Amongst the different nanoparticles to be employed, HGNs have the capacity to resonate in the near infrared region when irradiated by an appropriated laser (808 nm). By transforming this absorbed energy into heat, they are capable to produce locally induced hyperthermia. At this wavelength healthy tissues have a minimal light absorption being the effect restricted to the tissues containing HGNs. By placing HGNs inside MSCs, the recognition, excretion and immune response are minimized. We demonstrate that MSCs internalize HGNs and reach the tumors still containing HGNs. After laser treatment this loaded cells are able to eradicate tumoral cells in vitro and in vivo without significant toxicity. Also Ki67 expression, which is usually correlated with proliferation, is reduced after treatment. This approach enhances the effectiveness of the treatment when compared to just the enhanced permeation and retention effect (EPR) of the HGNs by themselves., Financial support from the EU thanks to the ERC Consolidator Grant program (ERC-2013-CoG-614715, NANOHEDONISM) is gratefully acknowledged.

The European wild rabbit ( Oryctolagus cuniculus) is a key prey species on the Iberian Peninsula, and several predator species that are at risk of extinction are dependent on them as prey. A new rabbit hemorrhagic disease (RHD) virus genotype (GI.2/RHDV2/b) emerged in 2010 and posed a threat to wild rabbit populations. During a survey aimed at investigating RHD epidemiology in wild rabbits, GI.2/RHDV2/b was detected by duplex real-time PCR in carcasses of one Mediterranean pine vole ( Microtus duodecimcostatus) and two white-toothed shrews ( Crocidura russula). Laboratory New Zealand white rabbits that were challenged with inocula obtained from the liver of the small mammals died showing RHD lesions, confirming the infectiousness of the isolates. Phylogenetic analysis of the VP60 gene nucleotide sequences showed complete homology between the isolates from the two small mammal species and a high degree of similarity, but not complete homology, to GI.2/RHDV2/b sequences from wild rabbits. The GI.2/RHDV2/b genotype has not been reported in species outside the order Lagomorpha.

Trabajo presentado en el Joint ESVP and ECVP Congress, celebrado en Cluj-Napoca (Rumanía), del 5 al 8 de septiembre de 2018, Introduction: In sheep, post-vaccination granulomas are a constant reaction after inoculation with vaccines containing aluminium-based adjuvants. These granulomas can be very persistent, sometimes lasting more than 15 months. Presence of multiple post-vaccination granulomas in the same animal can lead to chronic immune system stimulation and to the development of the ovine autoimmune/inflammatory syndrome induced by adjuvants (ASIA syndrome). Small ruminant lentiviruses (SRLV) infection is widespread and SRLV replicate in activated macrophages. The objective of this work is to report for the first time the active replication of SRLV in post-vaccination granulomas in sheep. Materials and Methods: Eleven adult, naturally SRLV-infected sheep showing clinical lentiviral arthritis were selected. Eight doses of commercial, aluminium-containing vaccines were inoculated in a period of 70 days to all animals. Granulomas were sampled and a study including pathology, cell culture (RT activity quantification), PCR and viral sequencing was carried out. SRLV circulating strains in each animal were also evaluated in peripheral blood mononuclear cells and spleen. Results: SRLV could be detected in post-vaccination granulomas in 5 (45.5%) sheep by cell culture and PCR. Viral sequences belonged to genotypes A and B and evinced compartmentalization according to the different tissues analysed including granulomas. Conclusions: Persistent granulomatous inflammatory reactions induced by vaccines allow in situ SRLV replication. As macrophages from these granulomas can be found in lymph nodes, SRLV may spread throughout the lymphatic system and likely increase the body viral load. The role of post-vaccination granulomas in the spread of sheep SRLV needs to be considered.

A 1-month-old Purebred Spanish Horse (PSH) foal presented with progressive hepatic failure culminating in death. Hepatic lesions were consistent with congenital hepatic fibrosis (CHF). Genetic studies in the PKHD1 gene in the affected foal revealed that it was heterozygous for the 2 previously described single-nucleotide polymorphisms (SNPs) linked to CHF in Swiss Franches-Montagnes (SFM) horses. In addition, 2 novel mutations were detected, the foal being homozygous for one of them and heterozygous for the other. Genetic studies in a healthy PSH population ( n = 35) showed a 3-fold higher genotypic frequency for PKHD1 SNP g.49,630,834G>A and a 5-fold higher genotypic frequency for PKHD1 SNP g.49,597,760A>T compared with those reported for SFM horses. SNPs in the PKHD1 gene in CHF-affected SFM horses might not fully explain the CHF observed in the PSH. Other mutations in the PKHD1 gene could play a more important role in the PSH.