Metagenomic Analysis of DNA Viruses with Targeted Sequence Capture of Canine Lobular Orbital Adenomas and Normal Conjunctiva.

Our study aims are: (1) to evaluate phenotypically normal canine conjunctival and orbital tissue and tissue from canine lobular orbital adenomas (CLOAs) for the presence of viral genomic material and (2) phylogenetically classify detected DNA viruses to determine if a DNA virus is associated with CLOAs. A total of 31 formalin fixed paraffin embedded CLOA tissue samples, 4 papillomas or sarcoid, and 10 fresh clinically normal conjunctival tissues were included in this study. Genomic DNA was isolated from all samples and sequencing libraries were prepared. The libraries were molecularly indexed and pooled and viral DNA was enriched via targeted sequence capture utilizing ViroCap. The libraries were sequenced on the Illumina HiSeq platform and compared to known viral DNA reference genomes to identify viral DNA. Carnivore parvovirus was identified in 6.4% and 20% of CLOA tissue and normal conjunctival samples, respectively. This study showed that conjunctival tissue from healthy dogs and CLOAs uncommonly harbor DNA viruses, and no DNA virus was associated with these tumors. Further studies are needed to evaluate the etiologic cause of CLOAs.

Papillomavirus DNA not detected in canine lobular orbital adenoma and normal conjunctival tissue.

BACKGROUND: Canine lobular orbital adenomas are benign tumors that arise from orbital glandular tissue and extend into the orbit, conjunctiva, and third eyelid. Surgical excision is challenging and recurrence rates are high following excision alone. Enucleation and exenteration reduces the likelihood of recurrence, but is a radical therapeutic option for an otherwise visual and comfortable eye. Human papillomavirus causes 4.5% of worldwide cancers in people and has been identified in up to 23% of benign salivary gland tumors. To date, the etiology of canine lobular orbital adenomas has not been established and it is reasonable to consider canine papillomaviruses as an associative agent with benign glandular tumors in dogs. Identification of the underlying etiology of these tumors may help establish treatment or preventative measures. The purpose of this study was to evaluate conjunctival and orbital tissue of phenotypically normal dogs and tissue from canine lobular orbital adenomas for the presence of papillomavirus DNA. RESULTS: Thirty seven canine lobular orbital adenoma samples (36 formalin fixed paraffin embedded (FFPE) tissue samples from 33 dogs and one freshly collected sample) were evaluated via polymerase chain reaction for the presence of papillomavirus DNA. Conjunctival tissue samples, from 10 dogs with normal ocular examinations, excised immediately following euthanasia, were used as phenotypically normal controls. Three FFPE and one freshly collected tissue samples previously confirmed to be positive for papillomavirus DNA were used as positive controls. PCR products verified positive controls. Papillomavirus DNA was not detected in fresh conjunctival tissue of the phenotypically normal control dogs or in samples of fresh or FFPE canine lobular orbital adenoma tissue. CONCLUSIONS: An association between papillomavirus and the development of canine lobular orbital adenomas is unlikely. Further research is needed to evaluate if other viruses play a role in the pathogenesis of canine lobular orbital adenomas.

Intravitreal implantation of TPP1-transduced stem cells delays retinal degeneration in canine CLN2 neuronal ceroid lipofuscinosis.

The CLN2 form of neuronal ceroid lipofuscinosis is a neurodegenerative disease that results from mutations in the TPP1 gene. Affected children exhibit progressive declines in most neurological functions including vision. Functional declines are accompanied by progressive brain and retinal atrophy. TPP1 encodes the soluble lysosomal enzyme tripeptidyl peptidase-1 (TPP1). Dachshunds with a TPP1 null mutation exhibit a disorder very similar to human CLN2 disease. Periodic infusion of recombinant TPP1 protein or a single injection of a TPP1 gene therapy vector into the cerebrospinal fluid of affected dogs significantly delays the onset and progression of neurological signs but does not slow vision loss or retinal degeneration. Studies were conducted to determine whether intravitreal implantation of autologous bone marrow derived stem cells transduced with a TPP1 expression construct would inhibit retinal degeneration in the canine model. A single injection of the transduced cells at an early stage in the disease progression substantially inhibited the development of disease-related retinal function deficits and structural changes. No adverse effects of the treatment were detected. These findings indicate that ex vivo gene therapy using autologous stem cells is an effective means of achieving sustained delivery of therapeutic compounds to tissues such as the retina for which systemic administration would be ineffective.

Surgical correction of congenital entropion in related Boer goat kids using a combination Hotz-Celsus and lateral eyelid wedge resection procedure.

Five related Boer goat kids (≤4 months of age) were presented to the University of Missouri, Veterinary Teaching Hospital (MU-VMTH) with epiphora and blepharospasm of several weeks duration and commencing prior to 1 month of age in all animals. Clinical examination confirmed euryblepharon and entropion bilaterally in two females and one male and unilaterally in two female kids. Deep stromal corneal ulceration was present in two eyes, and corneal granulation tissue and fibrosis were present in half (5/10) the affected eyes. A combination Hotz-Celsus and lateral eyelid wedge resection procedure was performed on all affected eyelids. Recheck examinations and long-term follow-up confirmed resolution of the entropion, preservation of normal eyelid conformation, and restoration of ocular comfort. Pedigree analysis ruled out sex-linked and autosomal dominant inheritance patterns; a specific mode of inheritance could not be determined. The Boer goat breed may be at increased risk for the development of entropion. This cases series represents the first report of entropion in the caprine species.

Local photodynamic therapy delays recurrence of equine periocular squamous cell carcinoma compared to cryotherapy.

OBJECTIVE: (i) To report the successful treatment of 10 cases of equine periocular squamous cell carcinoma (PSCC) with surgical excision and photodynamic therapy (PDT) using verteporfin. (ii) To evaluate time to first tumor recurrence between PDT-treated horses and horses treated with surgical excision and cryotherapy. METHODS: A total of 24 equine PSCC cases were included: group 1 (n = 14) had excision and cryotherapy (1993­2003), group 2 (n = 10), excision and local PDT (2006­2010). Evaluated data: signalment, treatment method, tumor location, size, and time to first recurrence. Groups were compared via chi-square test for categorical variables and Wilcoxon rank-sum test for numeric variables. Time to tumor recurrence was examined using Kaplan­Meier product-limit survival analysis. RESULTS: Of 24 cases, nine breeds were affected. Mean age at treatment in years: 14 (range 5­24) in group 1; 11 (range 8­18) in group 2. Median tumor size: 163 mm2 (range 20­625 mm2) in group 1; 195 mm2 (range 45­775 mm2) in group 2. Signalment, tumor laterality, and size were not significantly different between groups. Time to recurrence was significantly different between groups (Logrank test, P = 0.0006). In group 1, 11/14 horses had tumor regrowth with median time to recurrence in months: 10 (range 1­44). In group 2 (minimum follow-up of 25 months; range 25­50), no horse demonstrated tumor recurrence after one treatment with excision and PDT. CONCLUSIONS: This represents the first report of local PDT using verteporfin for treatment of equine PSCC. Following surgery, the likelihood of tumor recurrence was significantly reduced with local PDT compared with cryotherapy.

Long-term outcome of sudden acquired retinal degeneration syndrome in dogs.

OBJECTIVE: To investigate long-term outcomes and owner-perceived quality of life associated with sudden acquired retinal degeneration syndrome (SARDS) in dogs. DESIGN: Survey study. ANIMALS: 100 dogs with SARDS examined at 5 academic veterinary institutions from 2005 to 2010. PROCEDURES: The diagnosis was based on documented acute vision loss, normal results of ophthalmic examinations, and evaluation of extinguished bright-flash electroretinograms. Primary owners of affected dogs completed a questionnaire addressing outcome measures including vision, systemic signs, and perceived quality of life for their dogs. RESULTS: Age at diagnosis was significantly correlated with positive outcome measures; dogs in which SARDS was diagnosed at a younger age were more likely to have alleged partial vision and higher owner-perceived quality of life. Polyphagia was the only associated systemic sign found to increase in severity over time. Medical treatment was attempted in 22% of dogs; visual improvement was not detected in any. Thirty-seven percent of respondents reported an improved relationship with their dog after diagnosis, and 95% indicated they would discourage euthanasia of dogs with SARDS. CONCLUSIONS AND CLINICAL RELEVANCE: Blindness and concurrent systemic signs associated with SARDS appeared to persist indefinitely, but only polyphagia increased in severity over time. Most owners believed their pets had good quality of life and would discourage euthanasia of dogs with SARDS.

Surgical management of orbital nodular granulomatous episcleritis in a dog.

A 27 kg, 6-year-old, male castrated German shorthaired pointer presented to the University of Missouri, Veterinary Teaching Hospital with the complaint of progressive exophthalmia of 2 years duration optical density (OD). Lack of retropulsion OD was noted on physical examination. Anterior segment examination OU and fundic examination OS did not reveal any abnormalities. Examination of the fundus OD revealed focal scleral indentation of the inferior nasal globe. The indentation changed location with globe movement OD. MRI and CT scan revealed a well-circumscribed, approximately 2 cm in diameter mass located caudal and ventral to the affected globe that appeared to communicate with the nictitating membrane with absence of any bony involvement. A modified lateral orbitotomy was recommended and performed to remove the orbital mass and nictitating membrane en-bloc. Histopathology and immunohistochemistry of the mass confirmed a diagnosis of nodular granulomatous episcleritis (NGE). Postoperatively, the dog developed absolute keratoconjunctivitis sicca (KCS). Examples of primary episcleral inflammation in the dog include diffuse episcleritis, NGE, nodular fasciitis, fibrous histiocytoma, proliferative conjunctivitis/keratoconjunctivitis, pseudotumor, and Collie granuloma. The etiology of these episcleral inflammations is presumed to be immune mediated. To our knowledge, this is the first report of NGE affecting the orbital region of a dog. Development of absolute KCS resulting from excision of the nictitating membrane is also supported by this case.

Dacryops (lacrimal cyst) in three young labrador retrievers.

This case series constitutes a report of dacryops in multiple Labrador retrievers and the use of smooth-muscle actin immunostaining to confirm the lacrimal duct origins of the cyst wall. Three Labrador retrievers were presented with a history of a slowly enlarging mass adjacent to the left medial canthus. Ultrasonography of the masses revealed they were each spherical, thin-walled cystic structures. Aspiration cytology was performed in two cases revealing mixed inflammation and absence of detectable microorganisms. Dacryocystorhinography of the left nasolacrimal system performed in two cases revealed a normal nasolacrimal system that was closely associated, but not communicating with, the cystic mass in both cases. Surgical excision of all cysts was curative. Histopathology and positive immunohistochemical staining for smooth-muscle actin confirmed a diagnosis of dacryops in all cases.

Photodynamic therapy for the treatment of periocular squamous cell carcinoma in horses: a pilot study.

OBJECTIVE: Local photodynamic therapy (PDT) is a novel cancer therapy in veterinary ophthalmology. A prospective pilot study seeking to demonstrate proof of principle and safety for the treatment of equine periocular squamous cell carcinoma (PSCC) was therefore conducted. We hypothesized that surgical excision with adjunctive local PDT is an effective and safe treatment for equine PSCC. PROCEDURES: Nine horses (10 eyes) with PSCC were treated with surgical resection, local infiltration of resulting wound beds with 2-[1-hexyloxyethyl]-2-devinylpyropheophorbide-a (HPPH) and irradiation with 665-nm wavelength diode laser. Regular follow-up ophthalmic examinations were performed. RESULTS: Surgical resection and PDT yielded disease-free intervals of 25-68 months in our study horses as of January, 2008. These results were obtained following a single treatment in seven horses and two treatments in one horse. In one horse, carcinoma in situ developed 2.5 months after partial surgical excision and PDT, requiring local excision under standing sedation. CONCLUSIONS: Preliminary results suggest that surgical resection and adjunctive local PDT is a safe and effective novel treatment for PSCC in horses. More research is needed before PDT for the treatment of equine PSCC can be adequately compared with other current modalities. Important to future investigations regarding PDT, tumor recurrence rate, length of hospitalization, number of treatment episodes required to effect tumor remission, and total treatment costs should be examined in a controlled manner. Our present results and experiences suggest that this treatment may be useful in the treatment of equine PSCC.

Depletion of pulmonary intravascular macrophages inhibits acute lung inflammation.

Pulmonary intravascular macrophages (PIMs) are present in ruminants and horses. These species are highly sensitive to acute lung inflammation compared with non-PIM-containing species such as rats and humans. There is evidence that rats and humans may also recruit PIMs under certain conditions. We investigated precise contributions of PIMs to acute lung inflammation in a calf model. First, PIMs were recognized with a combination of in vivo phagocytic tracer Monastral blue and postembedding immunohistology with anti-CD68 monoclonal antibody. Second, gadolinium chloride depleted PIMs within 48 h of treatment (P < 0.05). Finally, PIMs contain TNF-alpha, and their depletion reduces cells positive for IL-8 (P < 0.05) and TNF-alpha (P < 0.05) and histopathological signs of acute lung inflammation in calves infected with Mannheimia hemolytica. The majority of IL-8-positive inflammatory cells in lung septa of infected calves were platelets. Platelets from normal cattle contained preformed IL-8 that was released upon in vitro exposure to thrombin (P < 0.05). These novel data show that PIMs, as the source of TNF-alpha, promote recruitment of inflammatory cells including IL-8-containing platelets to stimulate acute inflammation and pathology in lungs. These data may also be relevant to humans due to our ability to recruit PIMs.