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BACKGROUND: Nonexcisional tissue biopsies facilitate pre-operative confirmation of equine sarcoid yet fear of lesion deterioration currently limits its use in the diagnostic workup. OBJECTIVES: To evaluate the effect of a single punch biopsy on tumour growth dynamics [thickness, area, circumference, viral load (VL) and Visual Analog Scale (VAS)]. ANIMALS: Six client-owned horses with 11 sarcoids of various classification. MATERIALS AND METHODS: Growth dynamics were recorded on a weekly basis, 12 weeks pre- and 24 weeks post-biopsy. The effect of a single punch biopsy on growth dynamics was estimated by linear mixed-effect models. Spearman's rank correlation coefficient (SRCC) was calculated to investigate correlations with the number of weeks before and after the intervention. RESULTS: While statistically significant post-biopsy changes were recorded for all parameters except VL, no parameter was consistently increased after the biopsy intervention. In two of 11 sarcoids, the VL correlation pattern revealed significant strong correlations: sarcoid 6 (pre: r = -0.66, p < 0.05; post: r = 0.81, p < 0.001), increased VL; and sarcoid 8 (pre: r = 0.85, p < 0.001; post: r = 0.17, p > 0.05), no further increase after biopsy. CONCLUSIONS AND CLINICAL IMPORTANCE: Our results indicate that post-biopsy lesion deterioration is not a general concept that applies to all sarcoids, and both deterioration or improvement are possible outcomes over a 24-week period. Further clinical studies with a larger sample size are needed before a definitive conclusion can be made.
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Doenças dos Cavalos , Neoplasias Cutâneas , Humanos , Cavalos , Animais , Neoplasias Cutâneas/veterinária , Projetos Piloto , Pele , Biópsia/veterináriaRESUMO
BACKGROUND: Bovine papillomavirus (BPV) types 1 and 2 play a central role in the etiology of the most common neoplasm in horses, the equine sarcoid. The unknown mechanism behind the unique variety in clinical presentation on the one hand and the host dependent clinical outcome of BPV-1 infection on the other hand indicate the involvement of additional factors. Earlier studies have reported the potential functional significance of intratypic sequence variants, along with the existence of sarcoid-sourced BPV variants. Therefore, intratypic sequence variation seems to be an important emerging viral factor. This study aimed to give a broad insight in sarcoid-sourced BPV variation and explore its potential association with disease presentation. METHODS: In order to do this, a nanopore sequencing approach was successfully optimized for screening a wide spectrum of clinical samples. Specimens of each tumour were initially screened for BPV-1/-2 by quantitative real-time PCR. A custom-designed primer set was used on BPV-positive samples to amplify the complete viral genome in two multiplex PCR reactions, resulting in a set of overlapping amplicons. For phylogenetic analysis, separate alignments were made of all available complete genome sequences for BPV-1/-2. The resulting alignments were used to infer Bayesian phylogenetic trees. RESULTS: We found substantial genetic variation among sarcoid-derived BPV-1, although this variation could not be linked to disease severity. Several of the BPV-1 genomes had multiple major deletions. Remarkably, the majority of them cluster within the region coding for late viral genes. Together with the extensiveness (up to 603 nucleotides) of the described deletions, this suggests an altered function of L1/L2 in disease pathogenesis. CONCLUSIONS: By generating a significant amount of complete-length BPV genomes, we succeeded to introduce next-generation sequencing into veterinary research focusing on the equine sarcoid, thus facilitating the first report of both nanopore-based sequencing of complete sarcoid-sourced BPV-1/-2 and the simultaneous nanopore sequencing of multiple complete genomes originating from a single clinical sample.
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Papillomavirus Bovino 1 , Doenças dos Cavalos , Sequenciamento por Nanoporos , Nanoporos , Infecções por Papillomavirus , Neoplasias Cutâneas , Animais , Teorema de Bayes , Papillomavirus Bovino 1/genética , DNA Viral/genética , Genômica , Cavalos , Filogenia , Neoplasias Cutâneas/patologia , Neoplasias Cutâneas/veterináriaRESUMO
BACKGROUND: Donkeys are important throughout the world as work animals and occasionally as pets or a meat source. Most descriptions of skin disease in donkeys are reported in small case series, textbooks or review articles. HYPOTHESIS/OBJECTIVES: To document skin diseases and their prevalence in donkeys and to investigate predilections for the most common conditions. ANIMALS: Case populations at four veterinary schools totalling 156 donkeys. METHODS AND MATERIALS: A retrospective study was performed by searching computerized medical records, using the key word "donkey", at the School of Veterinary Medicine, University of California, Davis (UCD). Records of donkeys from the veterinary schools in Nantes, France; Utrecht, Netherlands and Ghent, Belgium were searched in a similar manner. The time periods included in the searches varied by institution. RESULTS: At UCD, 83 of 346 (24%) of donkeys had skin disease noted in their records. The most common diagnoses were insect bite hypersensitivity, sarcoid and habronemiasis. At Nantes, 36 of 144 (25%) had skin disease and the most common diagnoses were sarcoid and superficial pyoderma. At Utrecht 23 of 143 (16%) had skin disease and the most common diagnosis was dermatophytosis. At Ghent, 14 of 320 (4%) had skin disease and the most common diagnosis was sarcoid. CONCLUSIONS AND CLINICAL IMPORTANCE: Cutaneous conditions in donkeys are common. Age, sex and breed predisposition and the most common diagnoses varied with geographical location. Clinicians should include a dermatological examination regardless of the reason for presentation.
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Hipersensibilidade/veterinária , Dermatopatias/veterinária , Tinha/veterinária , Animais , Bélgica/epidemiologia , Equidae , França/epidemiologia , Hipersensibilidade/epidemiologia , Países Baixos/epidemiologia , Prevalência , Estudos Retrospectivos , Faculdades de Medicina Veterinária , Pele/patologia , Dermatopatias/epidemiologia , Tinha/epidemiologiaRESUMO
OBJECTIVE: To report the clinical features, outcomes, and prognostic factors associated with the surgical treatment of epiploic foramen entrapment (EFE). STUDY DESIGN: Retrospective study at a single referral hospital. ANIMALS: Horses (n = 142) undergoing surgery (n = 145) for EFE. METHODS: Preoperative, perioperative, and postoperative data of surgeries on horses that underwent exploratory laparotomy for EFE were obtained. The postoperative outcome was assessed by follow-up telephone calls with the owners/caregivers. Factors associated with postoperative reflux (POR), relaparotomy, hospital discharge, colic after hospital discharge, and survival after discharge were assessed. RESULTS: In total, 145 surgeries were performed on 142 horses (recurrence rate, 3%). Warmblood horses represented 85% of the horses that underwent surgery. Windsucking/crib-biting was confirmed in 60% of these surgery cases. Left-to-right entrapment was diagnosed in all horses. Ileal involvement was recorded in 74% of the cases. Uncontrollable intraoperative hemorrhage was encountered in 6% of the surgeries. One hundred seven (74%) horses recovered from surgery, and 65% of those survived to discharge. The rate of survival to discharge of all surgeries was 48%. The median survival of the cases that were discharged exceeded 3193 days. Horses requiring intestinal resection were predisposed to POR, and those undergoing jejunoileostomy were more prone to POR than those undergoing jejunojejunostomy. Horses with POR were less likely to be discharged than those without POR, and those that underwent resection had shorter life expectancy after hospital discharge than those that did not undergo resection. CONCLUSION: Surgical treatment of EFE was associated with high morbidity and mortality, with recurrence in at least 3% of surviving horses. CLINICAL SIGNIFICANCE: Owners of horses with EFE should be informed of the guarded prognosis associated with current surgical treatment.
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Cólica/veterinária , Procedimentos Cirúrgicos do Sistema Digestório/veterinária , Doenças dos Cavalos/cirurgia , Obstrução Intestinal/veterinária , Intestino Delgado/cirurgia , Animais , Cólica/cirurgia , Procedimentos Cirúrgicos do Sistema Digestório/métodos , Feminino , Refluxo Gastroesofágico/cirurgia , Cavalos , Obstrução Intestinal/cirurgia , Laparotomia/veterinária , Masculino , Complicações Pós-Operatórias/veterinária , Período Pós-Operatório , Estudos RetrospectivosRESUMO
OBJECTIVE: To determine the required rate of a detomidine infusion (loading dose 5 µg kg-1; initial rate 12.5 µg kg-1 hour-1) added to a constant infusion of methadone (0.2 mg kg-1; 0.05 mg kg-1 hour-1) for sedation in standing horses and ponies undergoing elective surgeries with appropriate local anaesthetic techniques. STUDY DESIGN: Prospective, clinical study. ANIMALS: Adult, healthy, client-owned, non-food-producing horses or ponies sedated for elective standing surgeries longer than 45 minutes. METHODS: At baseline (in the stables before administration of sedative agents), at 10 minutes after sedation and every 5 minutes thereafter, ataxia, sedation and surgical condition were evaluated; each scored 0-3. These scores were used to adjust the detomidine administration rate using the Ghent Sedation Algorithm. A 10 cm visual analogue scale (VAS) was used by the main surgeon at the end of the procedure to evaluate the surgical conditions. Heart rate, systolic arterial pressure and respiratory frequency were also recorded at each time point. For statistical analysis, anova for normal, Kruskal-Wallis H-test for non-normal variables, and Mann-Whitney U test for VAS were used. RESULTS: From the 42 horses/ponies included in this study, 28 underwent dental procedures and 14 other types of procedures. Overall, dental procedures required higher mean detomidine rates compared with other types of surgeries (16.9 ± 4.5 versus 9.0 ± 1.9 µg kg-1 hour-1) (p < 0.001). Dental procedures were assigned similar VAS scores, median (range), of 7.8 (5.8-10) with other procedures, 8.7 (2.8-10). Cardiovascular changes were not clinically significant. No signs or behavioural changes of abdominal pain were observed postoperatively. CONCLUSIONS AND CLINICAL RELEVANCE: Satisfactory surgical conditions were achieved using a combination of detomidine and methadone infusions with locoregional anaesthesia, with no adverse effects. Dental procedures required higher detomidine dose rates compared with other surgeries.
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Agonistas de Receptores Adrenérgicos alfa 2/administração & dosagem , Anestesia/veterinária , Anestésicos Combinados/administração & dosagem , Cavalos/cirurgia , Hipnóticos e Sedativos/administração & dosagem , Imidazóis/administração & dosagem , Metadona/administração & dosagem , Anestesia Dentária/veterinária , Animais , Infusões Intravenosas/veterinária , Estudos ProspectivosRESUMO
OBJECTIVE: To determine the incidence of infection and associated risk factors, after elective arthroscopy. STUDY DESIGN: Retrospective case study. ANIMALS: Horses (n=1079) undergoing elective arthroscopy. METHODS: Medical records of all horses that underwent elective arthroscopy between 2006 and 2013 were reviewed. Age, gender, breed, surgeon, number of joints operated, total anesthetic time, perioperative antimicrobial administration, and the presence and size of osteochondral fragments/subchondral lesions were recorded. For each operated joint, the development of postoperative infection (surgical site infection [SSI] and/or septic arthritis) and long-term outcome (>6 months) were recorded. Multivariate logistic regression was used to test for association between the independent variables and the dependent outcomes. RESULTS: A total of 1741 joints in 1079 horses underwent arthroscopy. SSI without septic arthritis occurred in 1 fetlock joint (0.14%), 1 tibiotarsal joint (0.19%), and 6 femoropatellar joints (1.67%). Thirteen joints (0.75%) were diagnosed with septic arthritis, including 1 fetlock joint (0.14%), 4 tibiotarsal joints (0.74%), and 8 femoropatellar joints (2.23%). The probability of postoperative SSI was higher when large lesions (>40 mm long) were treated, compared to medium (20-40 mm, P = .005) and small (<20 mm, P < .001) lesions. SSI was a significant risk factor for the development of septic arthritis (P < .001). Although age did not affect the incidence of SSI, increasing age was associated with a lower rate of septic arthritis rate (P = .028). CONCLUSION: Septic arthritis after elective arthroscopy was more likely in the presence of SSI and younger age. Horses with large lesions were at risk for SSI, which translated into a higher incidence of postoperative septic arthritis after femoropatellar arthroscopy.
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Artrite Infecciosa/veterinária , Artroscopia/veterinária , Procedimentos Cirúrgicos Eletivos/veterinária , Doenças dos Cavalos/etiologia , Infecção da Ferida Cirúrgica/veterinária , Animais , Antibacterianos/uso terapêutico , Artrite Infecciosa/etiologia , Artroscopia/efeitos adversos , Procedimentos Cirúrgicos Eletivos/efeitos adversos , Feminino , Doenças dos Cavalos/tratamento farmacológico , Cavalos , Incidência , Modelos Logísticos , Masculino , Complicações Pós-Operatórias/veterinária , Estudos Retrospectivos , Fatores de Risco , Infecção da Ferida Cirúrgica/etiologiaRESUMO
OBJECTIVE: To describe the anatomy of the entry to the equine esophagus (vestibulum esophagi) and to assess the risk of penetrating its adventitia and/or lumen during laryngoplasty. STUDY DESIGN: Ex vivo cadaveric study. SAMPLE POPULATION: Five isolated equine larynges and 39 equine head and neck specimens. METHODS: The anatomy of the vestibulum esophagi was studied by dissection of 5 cadaver specimens. Then, a bilateral laryngoplasty was performed, including 5 suture placements through the muscular processes, caudal, rostral, and sagittal, with straight and curved needles. Two of the 3 surgeons performing the implantations were unaware of the goals of the study. Suture positions and iatrogenic trauma to the lumen and/or adventitia of the vestibulum esophagi were identified during dissection of the specimens. Risk factors for penetrating the adventitia were evaluated with a multivariate regression model. RESULTS: The vestibulum esophagi spans between both wings of the thyroid cartilage over the entire width of the larynx, covering the rostral spine (arcuate crest) of the arytenoid cartilages. It is covered by the thyropharyngeus and cricopharyngeus muscles. Masked surgeons were associated with a significantly higher number of adventitia penetrations (72%) compared to the nonmasked surgeon (9%). The lumen of the vestibulum esophagi was penetrated in 4.6% of suture placements and only by the 2 masked surgeons. CONCLUSION: Penetration of the adventitia was more common when surgeons were unaware of the anatomical extent of the vestibulum esophagi. CLINICAL SIGNIFICANCE: Anatomical knowledge of the extent of the vestibulum esophagi reduces the risk of penetrating its lumen or adventitia during suture placement on the muscular process of the arytenoid cartilage.
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Esôfago/anatomia & histologia , Cavalos/anatomia & histologia , Laringoplastia/veterinária , Animais , Cartilagem Aritenoide/cirurgia , Cadáver , Esôfago/cirurgia , Laringe/cirurgiaRESUMO
BACKGROUND: Previous studies mention the use of topical acyclovir for the treatment of equine sarcoids. Success rates vary and since the bovine papillomavirus (BPV) lacks the presence of a kinase necessary to activate acyclovir, there is no proof of its activity against equine sarcoids. RESULTS: Twenty-four equine sarcoids were topically treated with acyclovir cream and 25 with a placebo. Both creams were applied twice daily during 6 months. Before the start of the treatment and further on a monthly basis, photographs and swabs were obtained. On the photographs, sarcoid diameter and surface area were measured and verrucosity of the tumours was quantified using a visual analog scale (VAS). The swabs were analysed by PCR for the presence of BPV DNA and positivity rates were calculated as the number of positive swabs divided by the total number of swabs for each treatment group at each time point. Success rates were not significantly different between both treatment groups. There was also no significant effect of treatment on sarcoid diameter, surface area or VAS score. For the swabs, a significantly higher BPV positivity rate was found for acyclovir treated tumours compared to placebo treated sarcoids only after 1 month of treatment and not at other time points. CONCLUSIONS: None of the results indicate that treatment with acyclovir yields any better results compared to placebo treatment.
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Aciclovir/uso terapêutico , Antineoplásicos/uso terapêutico , Antivirais/uso terapêutico , Doenças dos Cavalos/tratamento farmacológico , Neoplasias Cutâneas/veterinária , Aciclovir/administração & dosagem , Administração Tópica , Animais , Antineoplásicos/administração & dosagem , Antivirais/administração & dosagem , Método Duplo-Cego , Cavalos , Placebos , Creme para a Pele , Neoplasias Cutâneas/tratamento farmacológicoRESUMO
Background: Potential synovial penetration following palmar digital nerve blocks has not been investigated. Objectives: To evaluate the proximity of needles placed for palmar digital nerve blocks to nearby synovial structures using computed tomography (CT). Study design: Descriptive observational study. Methods: In 18 cadaver forelimbs, sequential injection of the navicular bursa (NB), distal interphalangeal joint (DIPJ) and digital flexor tendon sheath (DFTS) was performed using 3, 5 and 10 mL diluted radiodense contrast medium, respectively. After each synovial injection, 25 gage needles were placed over the palmar digital nerves at the proximal aspect of the ungular cartilages (distal injections) and 1 cm further proximally (proximal injections), and CT examination was performed. Subsequently, needles were removed, and the synovial structures further distended with the same volume as for the first injection. Perineural needle placement and image acquisition were repeated. The distance between the needle tip and adjacent synovial structures was measured (mm) in reconstructed images. Results were analyzed in separate general linear mixed models, to determine the effect of needle position and synovial distension on the distance from the tip of the needle to the NB, DFTS and DIPJ. Results: Synovial penetration was confirmed following 12/420 (3%) needle placements (NB n = 5, 1 after proximal and 4 after distal injections; DIPJ n = 2, DFTS n = 2, NB or DIPJ n = 3, all after distal injections). The mean distance from the needle tip to the NB and DIPJ was significantly smaller after the second distension (NB: p = 0.025; DIPJ: p < 0.001) and with the distal needle placements (NB: p < 0.001; DIPJ: p < 0.001). For the DFTS, the distance from the needle tip was significantly smaller with the proximal needle placements (p = 0.001). Main limitations: Ex-vivo study. Conclusion: There is a small risk of synovial penetration when performing palmar digital nerve blocks, especially when distension of adjacent synovial structures is present.
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This retrospective study aimed to assess the incidence of hypotension and the subsequent administration of dobutamine in horses anesthetized with isoflurane and romifidine during elective surgery. Time from induction of anaesthesia to administration of dobutamine was registered, as well as the time and dose needed to restore mean arterial pressure (MAP) ≥ 70 mmHg. Additionally, the influence of patient and anaesthesia related parameters on the need for dobutamine supplementation was evaluated. In total, 118 horses were included in this retrospective study. Dobutamine was administered to effect when MAP<70 mmHg. Data registered: patient weight, acepromazine premedication, body position, administration of intraoperative ketamine bolus, locoregional anaesthesia, mechanical ventilation, duration of anaesthesia, dose and duration of dobutamine administration, heart rate, MAP before dobutamine administration, MAP and time required to increase MAP≥70 mmHg. Dobutamine infusion was needed in 54.2% of the horses 30 ± 17 min after isoflurane-romifidine anaesthesia started. Dobutamine 0.55 ± 0.18 µg kg-1 min-1 achieved a MAP≥70 mmHg in 12 ± 8 min. Duration of dobutamine infusion was 56 ± 37 min. An univariable logistic regression showed a significant association between dobutamine and acepromazine administration (p = 0.01; OR = 3.43), anaesthesia time (p = 0.02; OR = 2.41) and dorsal recumbency (p < 0.001; OR = 8.40). In a multivariable logistic regression, only dorsal recumbency significantly increased the need for dobutamine supplementation (p < 0.001; OR = 7.70). There was no significant association between patient weight (p = 0.11; OR = 1), locoregional anaesthesia (p = 0.07; OR = 0.47), administration of a ketamine bolus (p = 0.95; OR = 0.98) or volume controlled ventilation (p = 0.94; OR = 1.04) and dobutamine administration. Low doses of dobutamine were suitable to restore MAP above 70 mmHg within a limited time period. Only dorsal recumbency increased the need of dobutamine administration.
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Anestesia , Anestésicos Inalatórios , Imidazóis , Isoflurano , Ketamina , Cavalos , Animais , Isoflurano/farmacologia , Dobutamina/farmacologia , Ketamina/farmacologia , Anestésicos Inalatórios/farmacologia , Acepromazina , Estudos Retrospectivos , Pressão Sanguínea , Anestesia/veterináriaRESUMO
Bovine papillomavirus (BPV) types 1 and 2 are causally associated with equine sarcoid, the most common mesenchymal neoplasm of horses, but the viral load (VL) differs between lesions. Sensitive and accurate BPV detection and quantification is essential for clinicians to confirm clinical suspicion, as well as in research settings for stratifying these skin lesions. Due to the limitations of histopathology in sarcoid diagnosis, PCR screening of superficial swabs constitutes the principal sampling method for BPV detection. This study aimed to investigate the ability of superficial swabs and fine-needle aspirates (FNA) to accurately detect the VL in equine sarcoids, considering the main clinical types: occult, nodular, verrucous and fibroblastic. Superficial swabs and FNAs from a series of sarcoid-affected horses were tested in parallel for BPV DNA quantification. Quantitative real-time PCR screening of postoperative tissue biopsies served as reference standard for the accuracy assessment of the viral titters. Our results indicate that VL is not a predictor of the clinical type. Student's t-test results gave evidence of a significant difference between both sample methods (P < 0.001) with FNA giving the best approximation of the actual VL (P < 0.01). In contrast to superficial swabs, the reference standard correlated moderately with FNA in general (P < 0.05; r = 0.39) and strongly with FNA results within the occult sarcoid group (P < 0.05; r = 0.59). In conclusion, the correlation of FNA with the reference standard was strong enough to suggest this is the preferred method for quantifying VL in sarcoids.
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Papillomavirus Bovino 1 , Doenças dos Cavalos , Neoplasias , Infecções por Papillomavirus , Sarcoidose , Dermatopatias , Neoplasias Cutâneas , Cavalos/genética , Animais , Carga Viral/veterinária , Infecções por Papillomavirus/diagnóstico , Infecções por Papillomavirus/veterinária , DNA Viral/análise , Dermatopatias/veterinária , Neoplasias/veterinária , Sarcoidose/diagnóstico , Sarcoidose/veterinária , Reação em Cadeia da Polimerase em Tempo Real/veterinária , Doenças dos Cavalos/diagnóstico , Neoplasias Cutâneas/diagnóstico , Neoplasias Cutâneas/veterinária , Papillomavirus Bovino 1/genéticaRESUMO
Sarcoids are the most common equine skin tumours Although they do not metastasize, they can be locally aggressive and cause significant clinical symptoms in affected horses. Despite being common, very little is known about the host immune response and the biological mechanisms underlying persistence and recurrence of equine sarcoids. The latter reflects the need for further research in this field. This in-vitro study used sarcoid explants from horses with naturally occurring sarcoids (n = 12) to evaluate the induction of a humoral immune response directed against equine sarcoid-derived bovine papilloma-virus (BPV)- 1 infected fibroblasts using a flow cytometric crossmatch assay. The presence of antibodies against exogenous bovine serum albumin (BSA) and fibroblast-like mesenchymal stromal cells (MSCs) was also evaluated by ELISA and flow cytometry, respectively. The viral load in the sarcoid explants, the corresponding cultured sarcoid fibroblasts, and matched peripheral blood mononuclear cells (PBMCs) from affected horses were determined by quantitative BPV-1/- 2 PCR analysis. Antibodies against autologous sarcoid cells were present in six out of twelve sarcoid-affected horses. Serum from all horses showed cross reactivity with allogeneic sarcoid cells, while only a part reacted with BSA or MSCs. Screening of host PBMCs demonstrated the absence of BPV E1 nucleic acids. Statistical analysis revealed a significantly higher mean viral load in the parental sarcoid tissue compared to the low passage fibroblasts (P < 0.001). These results support the hypothesis that sarcoid-affected horses may develop antibodies recognizing tumour-specific antigens. In contrast to sarcoid explants, equine PBMCs do not seem to contain complete BPV genomes. These results provide a basis for future investigations on the clinical relevance of these antibodies.
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Doenças dos Cavalos , Sarcoidose , Dermatopatias , Neoplasias Cutâneas , Animais , Cavalos , Leucócitos Mononucleares , Neoplasias Cutâneas/veterinária , Dermatopatias/veterinária , Sarcoidose/veterinária , Fibroblastos , DNA ViralRESUMO
Equine sarcoids (EqS) are fibroblast-derived skin tumors associated with bovine papillomavirus 1 and 2 (BPV-1 and -2). Based on Southern blotting, the BPV-1 genome was not found to be integrated in the host cell genome, suggesting that EqS pathogenesis does not result from insertional mutagenesis. Hence, CRISPR/Cas9 implies an interesting tool for selectively targeting BPV-1 episomes or genetically anchored suspected host factors. To address this in a proof-of-concept study, we confirmed the exclusive episomal persistence of BPV-1 in EqS using targeted locus amplification (TLA). To investigate the CRISPR/Cas9-mediated editing of BPV-1 episomes, primary equine fibroblast cultures were established and characterized. In the EqS fibroblast cultures, CRISPR-mediated targeting of the episomal E5 and E6 oncogenes as well as the BPV-1 long control region was successful and resulted in a pronounced reduction of the BPV-1 load. Moreover, the deletion of the equine Vimentin (VIM), which is highly expressed in EqS, considerably decreased the number of BPV-1 episomes. Our results suggest CRISPR/Cas9-based gene targeting may serve as a tool to help further unravel the biology of EqS pathogenesis.
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Sistemas CRISPR-Cas , Neoplasias Cutâneas , Animais , Cavalos , Oncogenes , Fibroblastos , Marcação de GenesRESUMO
Although bovine papillomavirus (BPV)-induced equine sarcoids are often identified solely by clinical examination, confirmation of the clinical diagnosis is essential for correct treatment selection. Only few approaches are presently available for this purpose. PCR-based screening for BPV in superficial swabs is widely-used to support clinical suspicion. While this method effectively detects sarcoid involvement in ulcerated lesions, sensitivity is lower in tumors with intact epithelium. This cross-sectional study compared the diagnostic characteristics of superficial swabs and fine-needle aspirates (FNA) with the aim to validate FNA as an alternative sampling method to detect BPV in suspect lesions. Among 63 lesions confirmed as sarcoids from 58 horses, compared to swabs FNA detected a greater proportion of BPV positives in general (swab, 70 %,;95 % confidence intervals [CI], 58.5 %-81.2 %; FNA, 98 %, 95 % CI, 95.3 %-100 %; P = 0.0001) and among a non-ulcerated subgroup (swab, 63 %, 95 % CI, 50.4 %-76.6 %; FNA, 98 %, 95 % CI, 91.4 %-100 %; P = 0.0001). Furthermore, the sensitivity, as well as the negative predictive value and accuracy of FNA for matched samples from 58 horses were superior to surface swabbing for the group of all horses and the subgroup that included only lesions with an intact epidermis (n = 48), with differences ranging from 23 % (95 % CI, 11.4 %-34.6 %) to 52 % (95 % CI, 25.9 %-78.1 %). Other advantages of FNA identified were a higher chance of adequate sampling and a reduced risk of unwanted detection of superficial contamination or latent BPV in keratinocytes. The ability to consistently detect BPV in all clinical lesion types indicates that FNA shows promise as a valid diagnostic tool to improve the consistency and quality of the diagnostic workup of equine sarcoids.
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Introduction: Enzootic pneumonia still causes major economic losses to the intensive pig production. Vaccination against its primary pathogen, Mycoplasma hyopneumoniae, is carried out worldwide to control the disease and minimize clinical signs and performance losses. Nonetheless, the effects of both infection with, and vaccination against Mycoplasma hyopneumoniae on the innate and adaptive immune responses remain largely unknown. Therefore, we conducted a study in which piglets were injected once with a commercial bacterin V1 or V2, or the adjuvant of V1 (A) to investigate their effect on local, innate and adaptive immune responses. Methods: Three weeks after vaccination, piglets were challenge infected with M. hyopneumoniae and euthanized four weeks later to assess vaccine efficacy via macroscopic and microscopic evaluation of lung lesions. Blood and broncho-alveolar lavage fluid (BAL) samples were collected to measure antibody responses, cellular immunity, BAL cytokine levels and BAL M. hyopneumoniae DNA load as well as cytokine secretion by monocytes. Results: After vaccination, proliferation of antigen-specific CD3+ T cells and a higher percentage of TNF-α+ CD8+, and TNF-α+ and TNF-α+IFN-γ+ CD4+CD8+ T cells was seen in V1, while proliferation of or a significant increase in cytokine production by different T cell subsets could not be observed for animals from V2. Interestingly, LPS-stimulated blood monocytes from V1 and A secreted less IL-10 on D7. After challenge, higher levels of IgA, more IL-10 and less IL-1ß was detected in BAL from V1, which was not observed in V2. Animals from A had significantly more IL-17A in BAL. The macroscopic lung lesion score and the M. hyopneumoniae DNA load at euthanasia was lower in V1, but the microscopic lung lesion score was lower in both vaccinated groups. Discussion: In conclusion, these results indicate that the two commercial bacterins induced different local and adaptive immune responses, that the adjuvant alone can reduce anti-inflammatory innate immune responses, and that both vaccines had a different efficacy to reduce Mycoplasma-like lung lesions and M. hyopneumoniae DNA load in the lung.
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Mycoplasma hyopneumoniae , Pneumonia Suína Micoplasmática , Suínos , Animais , Interleucina-10 , Fator de Necrose Tumoral alfa , Linfócitos T CD8-Positivos , Vacinas Bacterianas , Adjuvantes Imunológicos/farmacologia , Citocinas , Imunidade CelularRESUMO
BACKGROUND: Distal limb wounds in horses often show aberrant healing due to a slow inflammatory response. In human medicine, negative pressure wound therapy (NPWT) is used for the treatment of chronic wounds with a similar inflammatory response. OBJECTIVES: To compare the effect of NPWT to calcium alginate dressings on the healing of (non) contaminated equine distal limb wounds. STUDY DESIGN: Controlled experiment. METHODS: Circular wounds were created on the left and right dorsomedial metacarpus of 10 horses. In five horses, the wounds were contaminated with Staphylococcus aureus and Pseudomonas aeruginosa. In all horses, one limb was treated with NPWT, the other with calcium alginate dressings. Treatments were applied during nine days for noncontaminated wounds and six days for contaminated wounds. Noninvasive (clinical assessment, bacteriology swabs, thermographic images and wound dimensions) and invasive (biopsies for histology and growth factor analysis) measurements were taken regularly for 71 and 29 days respectively. Effects of selected parameters on continuous dependent variables were analysed using ANOVA, while for discrete dependent variables, logistic regression was applied. RESULTS: In noncontaminated wounds, there was significantly less wound retraction in the early healing stages when treated with NPWT (mean difference [95% CI] = 19.2% [13.3%-25.1%]; P = .005), although wound size was not significantly different between NPWT and control wounds at later healing stages. Noncontaminated control wounds had a significantly higher neutrophil influx (OR [95% CI] = 1.99 [1.49-2.66]; P < .001) and lower macrophage influx (OR [95% CI] = 0.75 [0.60-0.93]; P = .008) compared with NPWT-treated wounds. Bacterial load and the presence of growth factors did not differ between treatments in noncontaminated wounds. In contaminated wounds, no differences between treatments were observed in wound size, histological parameters, bacterial load or growth factor concentration. MAIN LIMITATIONS: Sample size is small. CONCLUSIONS: No long-term advantage was detected with NPWT compared with calcium alginate dressings in noncontaminated or contaminated equine distal limb wounds healing by second intention.
Assuntos
Tratamento de Ferimentos com Pressão Negativa , Alginatos , Animais , Bandagens/veterinária , Cavalos , Intenção , Tratamento de Ferimentos com Pressão Negativa/métodos , Tratamento de Ferimentos com Pressão Negativa/veterinária , CicatrizaçãoRESUMO
OBJECTIVE To evaluate outcomes following treatment of sarcoids in equids and to identify risk factors for treatment failure in these patients. DESIGN Retrospective case series. ANIMALS 230 equids with 614 sarcoids. PROCEDURES Records were searched to identify equids treated for ≥ 1 sarcoid between 2008 and 2013. A standardized protocol was used to determine treatment choice (electrosurgery, electrosurgery with intralesional placement of cisplatin-containing beads, topical administration of imiquimod or acyclovir, cryosurgery, bacillus Calmette-Guerin vaccine injection, or intralesional injection of platinum-containing drugs). Data regarding animal, tumor, treatment, and outcome variables were collected. Complete tumor regression without recurrence for ≥ 6 months was considered a successful outcome. Success rates were calculated; binary logistic regression analysis was used to identify risk factors for treatment failure and to compare effects of the 2 topical treatments. A χ(2) test was used to compare effects of the number of Bacillus Calmette-Guerin vaccine or cisplatin-containing drug injections on outcome. RESULTS The overall success rate was 460 of 614 (74.9%). Electrosurgical excision resulted in the highest treatment success rate (277/319 [86.8%]); odds of treatment failure were significantly greater for intralesional injection of platinum-containing drugs, cryosurgery, and topical acyclovir treatment. Odds of treatment failure were also significantly greater for sarcoids on equids with multiple tumors than for solitary lesions, and significantly lower for sarcoids on equids that received concurrent immunostimulating treatment for another sarcoid than for those on patients that did not receive such treatment. CONCLUSIONS AND CLINICAL RELEVANCE Selection bias for treatments was inherent to the study design; however, results may assist clinicians in selecting treatments and in determining prognosis for equids with sarcoids treated according to the described methods.
Assuntos
Doenças dos Cavalos/epidemiologia , Recidiva Local de Neoplasia/veterinária , Sarcoidose/veterinária , Neoplasias Cutâneas/veterinária , Animais , Bélgica/epidemiologia , Feminino , Doenças dos Cavalos/terapia , Cavalos , Masculino , Prontuários Médicos , Recidiva Local de Neoplasia/epidemiologia , Reoperação , Estudos Retrospectivos , Sarcoidose/epidemiologia , Neoplasias Cutâneas/epidemiologiaRESUMO
OBJECTIVE To compare antibacterial effects among 3 types of foam used with negative-pressure wound therapy (NPWT) in an ex vivo equine perfused wound model. SAMPLES Abdominal musculocutaneous flaps from 6 equine cadavers. PROCEDURES Each musculocutaneous flap was continuously perfused with saline (0.9% NaCl) solution. Four 5-cm circular wounds were created in each flap and contaminated with 106 CFUs of both Pseudomonas aeruginosa and methicillin-resistant Staphylococcus aureus (MRSA). After a 1-hour incubation period, 1 of 4 treatments (NPWT with silver-impregnated polyurethane foam [NPWT-AgPU], polyurethane foam [NPWT-PU], or polyvinyl alcohol foam [NPWT-PVA] or a nonadherent dressing containing polyhexamethylene biguanide without NPWT [control]) was randomly applied to each wound. An 8-mm punch biopsy specimen was obtained from each wound immediately before and at 6, 12, 18, and 24 hours after treatment application to determine the bacterial load for both P aeruginosa and MRSA. RESULTS The bacterial load of P aeruginosa for the NPWT-PVA treatment was significantly lower than that for the other 3 treatments at each sampling time after application, whereas the bacterial load for the NPWT-AgPU treatment was significantly lower than that for the NPWT-PU and control treatments at 12 hours after application. The bacterial load of MRSA for the NPWT-PVA treatment was significantly lower than that for the other 3 treatments at each sampling time after application. CONCLUSIONS AND CLINICAL RELEVANCE Results indicated that wounds treated with NPWT-PVA had the greatest decrease in bacterial load; however, the effect of that treatment on wound healing needs to be assessed in vivo.