Clinicopathological findings in cats with haws syndrome.

BACKGROUND: Haws syndrome is a relatively common yet poorly understood disorder in cats. METHODS: Ten cats with acute haws syndrome underwent ophthalmic and physical examinations, bloodwork, faecal analysis and pharmacological testing with 1% phenylephrine. The cats were discharged with broad-spectrum deworming (with or without metronidazole) as well as topical tetrahydrozoline. RESULTS: Cats were 17.6 ± 9.1 (2‒36) months old. All eyes had bilateral third eyelid protrusion and ptosis but no miosis or other ocular abnormalities. Diarrhoea was noted in four of 10 cats; otherwise, the cats were systemically healthy. A newly adopted kitten with haws syndrome and diarrhoea caused the condition to develop in the other three cats (from the same multicat household) within 4‒11 days. All cats tested negative for feline immunodeficiency virus and feline leukaemia virus. Serum amyloid A levels were elevated in two of eight tested cats. Tests for the presence of Giardia were positive in four of nine tested cats. Clinical signs transiently resolved within 20 minutes of the application of topical phenylephrine or tetrahydrozoline. Haws syndrome resolved in nine of the 10 cats within 37.7 ± 23 (12‒95) days but recurred in three of the nine cats 156‒182 days later. LIMITATIONS: The small sample size limits the conclusions that can be drawn. CONCLUSIONS: Haws syndrome occurs in generally healthy cats, with or without diarrhoea, and is characterised by a sympathetic neuropathy causing bilateral third eyelid protrusion and ptosis. Clinical signs are possibly triggered by contagious gastrointestinal pathogen(s) that presumably affect(s) the gut microbiota and influence(s) the gut‒brain axis.

Case report: Blindness associated with Learedius learedi trematode infection in a green sea turtle, Chelonia mydas, of the northern Red Sea.

Spirorchiid blood flukes are widespread in sea turtles, causing disease and mortality in their populations, with high prevalence in several ocean basins. Besides being leading parasitic causes of sea turtle strandings in several parts of the world, these infectious agents can cause endocarditis, vasculitis, thrombosis, miliary egg granulomas, and aneurysms, which ultimately may compromise the survival of green sea turtles. More severe cases may also result in multifocal granulomatous meningitis or pneumonia, both of which can be fatal. Herein, we report the first case of severe trematode infection, Caused by Learedius learedi, in a green sea turtle in the northern Red Sea; this infection is associated with bilateral blindness. Necropsy revealed multiple granulomas with intralesional trematode eggs in the optic nerve, eyes, spleen, heart, and lungs. The parasite was identified as Learedius learedi through specific primers of the ribosomal genome and COI sequences obtained from GenBank. Altogether, these findings emphasize the importance of recognizing the systemic nature of this particular fluke infection to ultimately protect the lives of these marine animals and ensure the sustainability of these species in the wild.

The 100 most-cited articles published in the Veterinary Ophthalmology journal between 1998 and 2022: A bibliometric study.

PURPOSE: The aim of this study was to bibliometrically analyze the 100 most-cited articles published in the Veterinary Ophthalmology (VO) journal. METHODS: Web of Science was searched for citations of VO articles published in 1998-2022. Tissue and species studied, and first and last author domicile and affiliation were recorded for the 100 most-cited articles and descriptively analyzed. RESULTS: Altogether, the 100 most-cited VO articles have cited a total of 5483 times. Most commonly, these were devoted to the cornea (23%), multiple tissues (19%), and glaucoma (16%). Studies on dogs (36%), horses (17%), and multiple species (15%) were most often cited. Most first/last authors were from the USA (n = 113), Brazil (n = 13), and France and Germany (n = 7 each), and most frequently affiliated with the University of Florida (n = 36), University of Wisconsin-Madison (n = 15), and Animal Health Trust, North Carolina State University, and Ohio State University (n = 6 each). KN Gelatt (n = 9), DE Brooks (n = 6), and FJ Ollivier and EO MacKay (n = 5 each) were the most frequent first or last authors. The greatest number of citations was for articles with KN Gelatt (n = 555), FJ Ollivier (n = 411), and DE Brooks (n = 372) as first or last authors. "The comparative morphology of the tapetum lucidum" by FJ Ollivier et al. (2002) is the most frequently cited article in VO history (n = 178). CONCLUSIONS: This study provides insight into the impact of publishing in VO and a more comprehensive understanding of trends and the most influential contributions to VO.

Retrospective evaluation of pre-surgical electroretinography results in a mixed-breed canine population presented for cataract removal surgery.

OBJECTIVE: Electroretinography (ERG) is used prior to cataract removal surgery to assess retinal function. We aimed to replicate and improve upon previous studies by performing a full ECVO protocol and by examining the retina post-surgery in all patients. ANIMALS STUDIED: One hundred twenty-seven eyes from 67 dogs were included in the study. PROCEDURES: A full ECVO protocol electroretinography, which includes extensive rod and cone analysis, was performed on all dogs presenting for cataract surgery. RESULTS: Our main findings were that amplitudes, but not implicit times of rod responses decreased with advanced cataracts. Amplitudes of the single flash rod and rod flicker responses were significantly lower in eyes with mature cataracts, and the former also decreased in hypermature cataracts. Cone flicker amplitude responses were also significantly lower in eyes with mature and hypermature cataracts. However, mixed single flash rod-cone and cone responses, with the exception of the mixed rod-cone a-wave amplitude in eyes with hypermature cataracts, were unaffected by cataract stage. The b-wave amplitude of the scotopic, mixed rod-cone, and photopic cone responses were affected by age and decreased by an average of 2.9, 7.5, and 1.5 µV/year, retrospectively (p < 0.01). CONCLUSIONS: Lower ERG amplitudes in canine cataract patients may result from aging or the presence of advanced cataracts and may not indicate the presence of retinal disease.

Treatment with chloroquine is retinotoxic in captive African penguins (Speniscus demersus). Attenuation and recovery of electroretinographic responses.

PURPOSE: To evaluate the effect of prophylactic anti-malarial chloroquine treatment, and its cessation, on electroretinographic (ERG) responses of captive African penguins. METHODS: A brief ERG protocol ("QuickRetCheck") was recorded under mesopic conditions with manual restraint and no sedation or pupil dilation. Birds were recorded on two separate occasions, first while being treated with a daily chloroquine dose of 10 mg/kg for 12 days (n = 15, treatment group) and second after 4 months without chloroquine treatment (n = 6, off-treatment group). Three birds were recorded on both occasions. Three other birds from the flock that died were studied histopathologically. RESULTS: Scotopic responses were unmeasurable in either recording and therefore were not analyzed. Mean a- and b-wave amplitudes of the mixed rod-cone responses to standard (3 cd·s/m2 ) and high (10 cd·s/m2 ) intensity flashes were higher in the off-treatment group. No difference in implicit times was observed. Sex, age, and number of previous chloroquine treatments did not affect ERG responses. Histopathology revealed Plasmodium spp.in the lungs, liver, and brain, but not in the eyes, of the necropsied birds, and there were no signs of retinitis or retinopathy. CONCLUSIONS: Daily chloroquine treatment was associated with attenuated ERG responses in penguins, which improve following cessation of treatment. Further work is warranted to establish a chloroquine dose that is efficacious yet has minimal adverse effects. Our results suggest that ERG responses of captive penguins undergoing ERG for any indication (such as prior to cataract surgery), must be evaluated in light of the birds' anti-malaria treatment status.

Atypical free-floating iridociliary pigmented epithelial cysts and secondary glaucoma in a caiman (Caiman latirostris).

We describe a case of chronic ocular trauma that resulted in fixed and free-floating, pigmented epithelial iridociliary cysts, inflammation, and secondary glaucoma in a caiman (Caiman latirostris). A 20- to 25-year-old male caiman was presented with phthisis bulbi in the right eye, and congested episcleral vessels, corneal leukoma, disorganized anterior chamber, multifocal anterior synechia, and elevated intraocular pressure in the left eye. Ocular ultrasound of the left eye revealed round structures dispersed in the anterior and posterior chambers and vitreous cavity. Bilateral enucleation was performed, and gross pathology of the left eye revealed multiple pigmented cysts attached to the iris and posterior corneal surface causing marked distortion of the anterior uvea, and free-floating in the vitreous cavity. Histopathology demonstrated heavily pigmented cystic structures of iridociliary epithelium origin carpeting the anterior segment surfaces and causing obstruction of the iridocorneal angles, leading to secondary glaucoma. To the authors' knowledge, this is the first report of iridociliary cysts in wildlife species.

The future of retinal gene therapy: evolving from subretinal to intravitreal vector delivery.

Inherited retinal degenerations are a leading and untreatbale cause of blindness, and as such they are targets for gene therapy. Numerous gene therapy treatments have progressed from laboratory research to clinical trails, and a pioneering gene therapy received the first ever FDA approval for treating patients. However, currently retinal gene therapy mostly involves subretinal injection of the therapeutic agent, which treats a limited area, entails retinal detachment and other potential complications, and requires general anesthesia with consequent risks, costs and prolonged recovery. Therefore there is great impetus to develop safer, less invasive and cheapter methods of gene delivery. A promising method is intravitreal injection, that does not cause retinal detachment, can lead to pan-retinal transduction and can be performed under local anesthesia in out-patient clinics. Intravitreally-injected vectors face several obstacles. First, the vector is diluted by the vitreous and has to overcome a long diffusion distance to the target cells. Second, the vector is exposed to the host's immune response, risking neutralization by pre-existing antibodies and triggering a stronger immune response to the injection. Third, the vector has to cross the inner limiting membrane which is both a physical and a biological barrier as it contains binding sites that could cause the vector's sequestration. Finally, in the target cell the vector is prone to proteasome degradation before delivering the transgene to the nucleus. Strategies to overcome these obstacles include modifications of the viral capsid, through rational design or directed evolution, which allow resistance to the immune system, enhancement of penetration through the inner limiting membrane or reduced degradation by intracellular proteasomes. Furthermore, physical and chemical manipulations of the inner limiting membrane and vitreous aim to improve vector penetration. Finally, compact non-viral vectors that can overcome the immunological, physical and anatomical and barriers have been developed. This paper reviews ongoing efforts to develop novel, safe and efficacious methods for intravitreal delivery of therapeutic genes for inherited retinal degenerations. To date, the most promising results are achieved in rodents with robust, pan-retinal transduction following intravitreal delivery. Trials in larger animal models demonstrate transduction mostly of inner retinal layers. Despite ongoing efforts, currently no intravitreally-injected vector has demonstrated outer retinal transduction efficacy comparable to that of subretinal delivery. Further work is warranted to test promising new viral and non-viral vectors on large animal models of inherited retinal degenerations. Positive results will pave the way to development of the next generation of treatments for inherited retinal degeneration.

Atypical ocular Chelonoid herpesvirus manifestations in a captive Loggerhead turtle (Caretta caretta).

A captive loggerhead turtle (Caretta caretta) of unknown sex, 3 years of age, presented with bilateral mucoid secretions, severe chemosis, conjunctival hyperemia, and globe retraction. The animal was evaluated ophthalmologically and systemically, and hematological, microbiological, and conjunctival cytological and biopsy samples were collected for complementary diagnosis. The histopathological examination showed amphophilic intranuclear inclusions associated with severe inflammatory infiltrate. The diagnosis of Chelonid alphaherpesvirus 5 (ChAHV 5) was confirmed with end point PCR. Following systemic treatment with L-lysine, acyclovir and vitamin A, the ocular signs resolved. No amphophilic intranuclear inclusions were seen in a follow-up biopsy 5 months later, and there has been no recurrence of clinical ophthalmic signs during a 4-year follow-up. It is suggested that ChAHV 5 be considered as a differential diagnosis in captive marine turtles that present for conjunctival disease other than fibropapillomatosis.

Evaluation of Photoreceptor Transduction Efficacy of Capsid-Modified Adeno-Associated Viral Vectors Following Intravitreal and Subretinal Delivery in Sheep.

Gene augmentation therapy based on subretinal delivery of adeno-associated viral (AAV) vectors is proving to be highly efficient in treating several inherited retinal degenerations. However, due to potential complications and drawbacks posed by subretinal injections, there is a great impetus to find alternative methods of delivering the desired genetic inserts to the retina. One such method is an intravitreal delivery of the vector. Our aim was to evaluate the efficacy of two capsid-modified vectors that are less susceptible to cellular degradation, AAV8 (doubleY-F) and AAV2 (quadY-F+T-V), as well as a third, chimeric vector AAV[max], to transduce photoreceptor cells following intravitreal injection in sheep. We further tested whether saturation of inner limiting membrane (ILM) viral binding sites using a nonmodified vector, before the intravitreal injection, would enhance the efficacy of photoreceptor transduction. Only AAV[max] resulted in moderate photoreceptor transduction following intravitreal injection. Intravitreal injection of the two other vectors did not result in photoreceptor transduction nor did the saturation of the ILM before the intravitreal injection. However, two of the vectors efficiently transduced photoreceptor cells following subretinal injection in positive control eyes. Previous trials with the same vectors in both murine and canine models resulted in robust and moderate transduction efficacy, respectively, of photoreceptors following intravitreal delivery, demonstrating the importance of utilizing as many animal models as possible when evaluating new strategies for retinal gene therapy. The successful photoreceptor transduction of AAV[max] injected intravitreally makes it a potential candidate for intravitreal delivery, but further trials are warranted to determine whether the transduction efficacy is sufficient for a clinical outcome.

Prognostic indicators and outcome in dogs undergoing temporary tarsorrhaphy following traumatic proptosis.

PURPOSE: Retrospectively evaluate prognostic indicators and outcome in dogs undergoing temporary tarsorrhaphy following traumatic proptosis. METHODS: Medical records (2004-2017) were reviewed for signalment, cause and duration of proptosis, and clinical findings on admission. The operating faculty member, post-operative medications, and ophthalmic findings at last recheck were recorded. Owners of dogs with blinded eyes were surveyed regarding final outcome and satisfaction. Data were analyzed using Fisher's exact, Likelihood ratio, and Mann-Whitney tests. RESULTS: At last recheck, 17.5 ± 7.3 (mean ± SD) days post-operatively, vision was present in 12/43 eyes (28%). Vision at last recheck was not correlated with breed, cause or duration of proptosis, or post-operative medications (P > .05), but was correlated with presence of direct and indirect pupillary light reflexes (PLRs) on admission (P = .001 and .02, respectively), and with assessment and surgery performed by veterinary ophthalmologists rather than surgery or emergency personnel (P = .015). Mean satisfaction scores (graded 0-10) of 22 owners contacted 59.6 ± 48.2 months after last recheck were 7.8 ± 2.6 (n = 6), 5.7 ± 4.9 (n = 3), and 8.8 ± 1.3 (n = 13) for dogs with phthisical eyes, eyes requiring long-term medications, and blind but otherwise normal eyes, respectively (P = .284). CONCLUSIONS: Contrary to previous studies, breed, and cause and duration of proptosis, did not significantly affect outcome. Testing of direct and indirect PLR is a simple and significant prognostic indicator. Patients assessed and operated by a veterinary ophthalmologist have better prognosis.

Analgesic effects of intraorbital insertion of an absorbable gelatin hemostatic sponge soaked with 1% ropivacaine solution following enucleation in dogs.

OBJECTIVE: To evaluate analgesic effects and complications associated with intraorbital insertion of an absorbable gelatin hemostatic sponge (AGHS) soaked with 1% ropivacaine solution following enucleation in dogs. ANIMALS: 20 client-owned dogs undergoing enucleation. PROCEDURES: Dogs were randomly assigned to receive an AGHS soaked with 1% ropivacaine solution (n = 10) or saline (0.9% NaCl) solution (control group; 10) inserted intraorbitally prior to skin closure following enucleation. Carprofen (2 mg/kg [0.9 mg/lb]) was administered SC once after orotracheal extubation and then PO twice a day for 5 days. During the postoperative recovery period, apparent pain level was scored at various points with a modified short-form Glasgow Composite Pain Scale (score range, 0 to 19), and methadone was administered for rescue analgesia if any score was ≥ 5. After dogs returned home, owners recorded their behavior and apparent pain level for the first 3 days following enucleation. RESULTS: At extubation, the median (range) pain score was significantly higher in the control group (8 [2 to 14]) versus the ropivacaine group (3 [1 to 7]). A greater proportion of dogs in the control group received methadone (7/10 vs 1/10) and had crying or attention-seeking behavior on the first day following enucleation (7/10 vs 1/10). No complications were observed in either group. CONCLUSIONS AND CLINICAL RELEVANCE: Addition of intraorbital insertion of a ropivacaine-soaked AGHS to the analgesic protocol for dogs undergoing enucleation provided better analgesia than was achieved without this treatment as measured immediately and the first day after surgery, with no noted adverse effects.

Can an in vivo imaging system be used to determine localization and biodistribution of AAV5-mediated gene expression following subretinal and intravitreal delivery in mice?

Recombinant adeno associated viruses (AAV) are the most commonly used vectors in animal model studies of gene therapy for retinal diseases. The ability of a vector to localize and remain in the target tissue, and in this manner to avoid off-target effects beyond the site of delivery, is critical to the efficacy and safety of the treatment. The in vivo imaging system (IVIS) is a non-invasive imaging tool used for detection and quantification of bioluminescence activity in rodents. Our aim was to investigate whether IVIS can detect localization and biodistribution of AAV5 vector in mice following subretinal (SR) and intravitreal (IVT) injections. AAV5 carrying firefly luciferase DNA under control of the ubiquitous cytomegalovirus (CMV) promoter was injected unilaterally IVT or SR (in the central or peripheral retina) of forty-one mice. Luciferase activity was tracked for up to 60 weeks in the longest surviving animals, using repeated (up to 12 times) IVIS bioluminescence imaging. Luciferase presence was also confirmed immunohistochemically (IHC) and by PCR in representative animals. In the SR group, IVIS readings demonstrated luciferase activity in all (32/32) eyes, and luciferase presence was confirmed by IHC (4/4 eyes) and PCR (12/12 eyes). In the IVT group, IVIS readings demonstrated luciferase activity in 7/9 eyes, and luciferase presence was confirmed by PCR in 5/5 eyes and by IHC (2/2 eyes). In two SR-injected animals (one each from the central and peripheral injection sites), PCR detected luciferase presence in the ipsilateral optic nerves, a finding that was not detected by IVIS or IHC. Our results show that when evaluating SR delivery, IVIS has a sensitivity and specificity of 100% compared with the gold standard PCR. When evaluating IVT delivery, IVIS has a sensitivity of 78% and specificity of 100%. These finding confirm the ability of IVIS to detect in-vivo localized expression of AAV following SR delivery in the retina up to 60 weeks post-treatment, using repeated imaging for longitudinal evaluation, without fading of the biological signal, thereby replacing the need for post mortem processing in order to confirm vector expression. However, IVIS is probably not sensitive enough, compared with genome detection, to demonstrate biodistribution to the optic nerve, as it could not detect luciferase activity in ipsilateral optic nerves following SR delivery in mice.

Six Years and Counting: Restoration of Photopic Retinal Function and Visual Behavior Following Gene Augmentation Therapy in a Sheep Model of CNGA3 Achromatopsia.

Achromatopsia causes severely reduced visual acuity, photoaversion, and inability to discern colors due to cone photoreceptor dysfunction. In 2010, we reported on day-blindness in sheep caused by a stop-codon mutation of the ovine CNGA3 gene and began gene augmentation therapy trials in this naturally occurring large animal model of CNGA3 achromatopsia. The purpose of this study was to evaluate long-term efficacy and safety results of treatment, findings that hold great relevance for clinical trials that started recently in CNGA3 achromatopsia patients. Nine day-blind sheep were available for long-term follow up. The right eye of each sheep was treated with a single subretinal injection of an Adeno-Associated Virus Type 5 (AAV5) vector carrying either a mouse (n = 4) or a human (n = 5) CNGA3 transgene under control of the 2.1-Kb red/green opsin promoter. The efficacy of treatment was assessed periodically with photopic maze tests and electroretinographic (ERG) recordings for as long as 74 months postoperatively. Safety was assessed by repeated ophthalmic examinations and scotopic ERG recordings. The retinas of three animals that died of unrelated causes >5 years post-treatment were studied histologically and immunohistochemically using anti-hCNGA3 and anti-red/green cone opsin antibodies. Passage time and number of collisions of treated sheep in the photopic maze test were significantly lower at all follow-up examinations as compared with pretreatment values (p = 0.0025 and p < 0.001, respectively). ERG Critical Flicker Fusion Frequency and flicker amplitudes at 30 and 40 Hz showed significant improvement following treatment (p < 0.0001) throughout the study. Ophthalmic examinations and rod ERG recordings showed no abnormalities in the treated eyes. Immunohistochemistry revealed the presence of CNGA3 protein in red/green opsin-positive cells (cones) of the treated eyes. Our results show significant, long-term improvement in cone function, demonstrating a robust rescue effect up to six years following a single treatment with a viral vector that provides episomal delivery of the transgene. This unique follow-up duration confirms the safe and stable nature of AAV5 gene therapy in the ovine achromatopsia model.

Electroretinography is a prognostic indicator for postoperative vision in dogs undergoing retinal reattachment surgery.

PURPOSE: To determine whether pre-operative electroretinography (ERG) predicts postoperative vision in dogs undergoing retinal reattachment surgery (RRS). METHODS: This 18-month prospective study recorded signalment, duration, cause, and extent of retinal detachment and pre-operative vision status. Rod and mixed rod-cone ERG responses were recorded prior to RRS. Referring veterinary ophthalmologists assessed vision 2 months postoperatively. RESULTS: Thirty dogs (40 affected eyes) aged 4 months to 12.1 years were included. The detachment extent was 150° -320° in 15 of 40 eyes, 360° in 24 of 40 eyes, and not recorded in one eye. Most dogs had a genetic predisposition for retinal detachment. Eight eyes of seven dogs had previous cataract surgery. Mean estimated duration of detachment prior to surgery was 24.5 ± 19.6 days. Pre-operatively, 34 of 40 eyes were blind, two of 40 eyes were sighted, and four of 40 eyes had severely diminished vision. Compared to normative ERG values in our clinics, pre-operative ERGs were classified as "normal" in five of 40 eyes, "attenuated" in seven of 40 eyes, and "flat" in 28 of 40 eyes. Following RRS, the retina was fully reattached in all operated eyes. Two-month postoperatively, 30 of 40 eyes had "normal" vision as defined by referring veterinary ophthalmologists, six of 40 eyes had "limited" or "diminished" vision and four of 40 eyes were blind. Normal vision was regained in 12 of 12 (100%) of eyes with normal or attenuated pre-operative ERG's, but only in 18 of 28 (64%) of eyes with flat pre-operative ERG 's (Linear-by-linear test, P = 0.029). CONCLUSIONS: A recordable pre-operative ERG, even if attenuated, is associated with return of vision in canine RRS patients, and is a favorable prognostic indicator.

Changes in Retinal Function and Cellular Remodeling Following Experimental Retinal Detachment in a Rabbit Model.

Purpose. To explore functional electroretinographic (ERG) changes and associated cellular remodeling following experimental retinal detachment in a rabbit model. Methods. Retinal detachment was created in ten rabbits by injecting 0.1 ml balanced salt solution under the retina. Fundus imaging was performed 0, 3, 7, 14, and 21 days postoperatively. ERGs were recorded pre- and 7 and 21 days postoperatively. Eyes were harvested on day 21 and evaluated immunohistochemically (IHC) for remodeling of second- and third-order neurons. Results. Retinal reattachment occurred within two weeks following surgery. No attenuation was observed in the photopic or scotopic a- and b-waves. A secondary wavefront on the descending slope of the scotopic b-wave was the only ERG result that was attenuated in detached retinas. IHC demonstrated anatomical changes in both ON and OFF bipolar cells. Bassoon staining was observed in the remodeled dendrites. Amacrine and horizontal cells did not alter, but Muller cells were clearly reactive with marked extension. Conclusion. Retinal detachment and reattachment were associated with functional and anatomical changes. Exploring the significance of the secondary scotopic wavefront and its association with the remodeling of 2nd- and 3rd-order neurons will shade more light on functional changes and recovery of the retina.

Safety and Efficacy Evaluation of rAAV2tYF-PR1.7-hCNGA3 Vector Delivered by Subretinal Injection in CNGA3 Mutant Achromatopsia Sheep.

Applied Genetic Technologies Corporation (AGTC) is developing a recombinant adeno-associated virus (rAAV) vector expressing the human CNGA3 gene designated AGTC-402 (rAAV2tYF-PR1.7-hCNGA3) for the treatment of achromatopsia, an inherited retinal disorder characterized by markedly reduced visual acuity, extreme light sensitivity, and absence of color discrimination. The results are herein reported of a study evaluating safety and efficacy of AGTC-402 in CNGA3-deficient sheep. Thirteen day-blind sheep divided into three groups of four or five animals each received a subretinal injection of an AAV vector expressing a CNGA3 gene in a volume of 500 µL in the right eye. Two groups (n = 9) received either a lower or higher dose of the AGTC-402 vector, and one efficacy control group (n = 4) received a vector similar in design to one previously shown to rescue cone photoreceptor responses in the day-blind sheep model (rAAV5-PR2.1-hCNGA3). The left eye of each animal received a subretinal injection of 500 µL of vehicle (n = 4) or was untreated (n = 9). Subretinal injections were generally well tolerated and not associated with systemic toxicity. Most animals had mild to moderate conjunctival hyperemia, chemosis, and subconjunctival hemorrhage immediately after surgery that generally resolved by postoperative day 7. Two animals treated with the higher dose of AGTC-402 and three of the efficacy control group animals had microscopic findings of outer retinal atrophy with or without inflammatory cells in the retina and choroid that were procedural and/or test-article related. All vector-treated eyes showed improved cone-mediated electroretinography responses with no change in rod-mediated electroretinography responses. Behavioral maze testing under photopic conditions showed significantly improved navigation times and reduced numbers of obstacle collisions in all vector-treated eyes compared to their contralateral control eyes or pre-dose results in the treated eyes. These results support the use of AGTC-402 in clinical studies in patients with achromatopsia caused by CNGA3 mutations, with careful evaluation for possible inflammatory and/or toxic effects.

Gene Augmentation Therapy for a Missense Substitution in the cGMP-Binding Domain of Ovine CNGA3 Gene Restores Vision in Day-Blind Sheep.

Purpose: Applying CNGA3 gene augmentation therapy to cure a novel causative mutation underlying achromatopsia (ACHM) in sheep. Methods: Impaired vision that spontaneously appeared in newborn lambs was characterized by behavioral, electroretinographic (ERG), and histologic techniques. Deep-sequencing reads of an affected lamb and an unaffected lamb were compared within conserved genomic regions orthologous to human genes involved in similar visual impairment. Observed nonsynonymous amino acid substitutions were classified by their deleteriousness score. The putative causative mutation was assessed by producing compound CNGA3 heterozygotes and applying gene augmentation therapy using the orthologous human cDNA. Results: Behavioral assessment revealed day blindness, and subsequent ERG examination showed attenuated photopic responses. Histologic and immunohistochemical examination of affected sheep eyes did not reveal degeneration, and cone photoreceptors expressing CNGA3 were present. Bioinformatics and sequencing analyses suggested a c.1618G>A, p.Gly540Ser substitution in the GMP-binding domain of CNGA3 as the causative mutation. This was confirmed by genetic concordance test and by genetic complementation experiment: All five compound CNGA3 heterozygotes, carrying both p.Arg236* and p.Gly540Ser mutations in CNGA3, were day-blind. Furthermore, subretinal delivery of the intact human CNGA3 gene using an adeno-associated viral vector (AAV) restored photopic vision in two affected p.Gly540Ser homozygous rams. Conclusions: The c.1618G>A, p.Gly540Ser substitution in CNGA3 was identified as the causative mutation for a novel form of ACHM in Awassi sheep. Gene augmentation therapy restored vision in the affected sheep. This novel mutation provides a large-animal model that is valid for most human CNGA3 ACHM patients; the majority of them carry missense rather than premature-termination mutations.

Conjunctival habronemiasis in a square-lipped rhinoceros (Ceratotherium simum).

A captive female square-lipped rhinoceros born in 1993 had been showing intermittent signs of bilateral conjunctivitis and conjunctival proliferation since 1998. Periodic improvement was noted, especially in winter, but overall the condition had deteriorated over the years. Treatment with various topical, intralesional, and systemic antibiotics and glucocorticosteroids was largely ineffective, as were repeated dewormings. No primary cause for these lesions was found in biopsies taken in 2000 and 2006, although a severe infiltrate of numerous eosinophils was observed in the latter. As the condition worsened, secondary corneal changes were noted, and eventually vision was lost due to proliferative conjunctival tissue. Aggressive resection of the proliferating tissue in 2013 restored vision and submitted biopsies yielded a diagnosis of severe allergic conjunctivitis, eosinophilic granuloma, and habronematid (Habronema or Draschia) larval infection. As no other rhinoceros in the herd was affected, including two calves born to the patient who were in close contact with their mother, it was concluded the presentation was most likely due to a hypersensitivity reaction to the dead or dying larvae. Fly repellent is now regularly applied around the eye of this rhinoceros, and a protective face mask has been fitted. Ongoing periodic relapses are treated with oral ivermectin, topical antibiotics, and steroids.

Polymethylmethacrylate orbital implants with interconnecting channels. A retrospective study following enucleation in dogs and cats.

PURPOSE: This study describes the use of polymethylmethacrylate implants with interconnecting channels (PIIC) to fill the orbit following enucleation in 31 dogs and 11 cats. METHOD: Seven channels were drilled into the implant. A central channel, running from the PIIC pole to its flat surface, was used to pass sutures anchoring the PIIC to the orbital fascia, minimizing the likelihood of extrusion. Six more channels allowed ingrowth of fibrovascular tissue into the PIIC, which reduces the risk of extrusion. Patients were evaluated 3, 10, and 30 days postoperatively, with 15 of 42 patients followed for 6-20 months. Ultrasound examination was performed in seven patients 15-510 days postoperatively, and two implants were studied histopathologically. RESULTS: Implants were well tolerated and prevented skin concavity, leading to good cosmetic results. Blood flow was imaged in vessels growing into the channels, and histopathologically a collagenous, fibrovascular capsule was seen surrounding the PIIC and invading its channels. CONCLUSIONS: PIICs are a safe and cosmetic solution to anophthalmic orbits, allowing ingrowth of blood vessels and fibrovascular tissue.

The effects of topical parasympatholytic drugs on pupil diameter and intraocular pressure in healthy dogs treated with 0.005% latanoprost.

PURPOSE: Prostaglandin analogs induce miosis and lower intraocular pressure (IOP). As pupils of latanoprost-treated eyes may have to be dilated for ophthalmoscopy or intraocular surgery, we studied whether 0.5% tropicamide or 1% atropine alter the effects of 0.005% latanoprost on pupil diameter (PD) and IOP in healthy dogs. METHODS: IOP and PD were measured hourly, 8 AM-4 PM, with the right and left eyes serving as control (CE) and treated (TE) eyes, respectively. Measurements were conducted in ten Labrador retrievers with one-week washout: (i) baseline values, (ii) latanoprost at 8 AM, (iii) tropicamide at 8 AM, (iv) latanoprost at 8 AM and tropicamide at 11 AM, and (v) latanoprost at 8 AM and atropine at 11 AM (n = 4). RESULTS: At 4 PM, TE PD was 5.88 ± 0.59, 3.62 ± 0.66, 6.33 ± 1.00, 5.42 ± 0.57, and 8.12 ± 1.24 mm in sessions 1-5, respectively. TE PD was significantly different between treatment sessions 2, 4, and 5 (P = 0.018, Friedman), being most mydriatic in session 5. At 4 PM, TE IOP was 11.27 ± 2.07, 7.10 ± 1.07, 11.1 ± 2.21, 7.70 ± 1.85, and 8.87 ± 1.42 mm Hg in sessions 1-5, respectively, with no differences between treatment sessions 2, 4, and 5 (P = 0.105, Friedman). CONCLUSIONS: Tropicamide and atropine counteracted latanoprost's miotic effect, with atropine causing significantly larger mydriasis, sufficient for indirect ophthalmoscopy. Neither drug counteracted the hypotensive effect of latanoprost during this study period. Further studies are necessary to evaluate the potential risks in glaucomatous dogs.