Achromatopsia is a genetic disorder of cones, and one of the most common forms is a channelopathy caused by mutations in the β-subunit, CNGB3, of the cone cyclic nucleotide-gated (CNG) channel. Recombinant adeno-associated virus of serotype 5 (rAAV5)-mediated gene transfer of human CNGB3 cDNA to mutant dog cones results in functional and structural rescue in dogs <0.5 years of age, but treatment is minimally effective in dogs >1 year. We now test a new therapeutic concept by combining gene therapy with the administration of ciliary neurotrophic factor (CNTF). Intravitreal CNTF causes transient dedifferentiation of photoreceptors, a process called deconstruction, whereby visual cells become immature with short outer segments, and decreased retinal function and gene expression that subsequently return to normal. Cone function was successfully rescued in all mutant dogs treated between 14 and 42 months of age with this strategy. CNTF-mediated deconstruction and regeneration of the photoreceptor outer segments prepares the mutant cones optimally for gene augmentation therapy.

Cyclic nucleotide-gated (CNG) ion channels are key mediators underlying signal transduction in retinal and olfactory receptors. Genetic defects in CNGA3 and CNGB3, encoding two structurally related subunits of cone CNG channels, lead to achromatopsia (ACHM). ACHM is a congenital, autosomal recessive retinal disorder that manifests by cone photoreceptor dysfunction, severely reduced visual acuity, impaired or complete color blindness and photophobia. Here, we report the first canine models for CNGA3-associated channelopathy caused by R424W or V644del mutations in the canine CNGA3 ortholog that accurately mimic the clinical and molecular features of human CNGA3-associated ACHM. These two spontaneous mutations exposed CNGA3 residues essential for the preservation of channel function and biogenesis. The CNGA3-R424W results in complete loss of cone function in vivo and channel activity confirmed by in vitro electrophysiology. Structural modeling and molecular dynamics (MD) simulations revealed R424-E306 salt bridge formation and its disruption with the R424W mutant. Reversal of charges in a CNGA3-R424E-E306R double mutant channel rescued cGMP-activated currents uncovering new insights into channel gating. The CNGA3-V644del affects the C-terminal leucine zipper (CLZ) domain destabilizing intersubunit interactions of the coiled-coil complex in the MD simulations; the in vitro experiments showed incompetent trimeric CNGA3 subunit assembly consistent with abnormal biogenesis of in vivo channels. These newly characterized large animal models not only provide a valuable system for studying cone-specific CNG channel function in health and disease, but also represent prime candidates for proof-of-concept studies of CNGA3 gene replacement therapy for ACHM patients.

Gallbladders and rectal contents were collected from cattle (n ‫؍‬ 933) at slaughter to determine whether the gallbladder harbors Escherichia coli O157:H7. Both gallbladder mucosal swabs and homogenized mucosal tissues were used for isolation. Only five gallbladders (0.54%) were positive for E. coli O157:H7. Fecal prevalence averaged 7.1%; however, none of the cattle that had E. coli O157:H7 in the gallbladder was positive for E. coli O157:H7 in feces. Therefore, the gallbladder does not appear to be a common site of colonization for E. coli O157:H7 in beef cattle.Escherichia coli O157:H7 is a major food-borne pathogen in the United States. Cattle are the main reservoir for E. coli O157:H7, which colonizes the gastrointestinal tract and is shed in the feces. Previous studies have documented that prevalence and shedding patterns of E. coli O157:H7 in the feces of beef cattle are highly variable (1, 4, 5). The prevalence ranges from 2 to 3% to as high as 80% of cattle sampled (1, 4). The shedding patterns in cattle vary from transient, in which animals shed the organism only for a few days, to persistent, in which shedding may last for an extended period of time (1,4,12). The persistent shedding is most likely due to the organism colonizing a part of the gastrointestinal tract, such as the mucosa of the rectoanal junction (11). Previous studies have shown that the gallbladder may be a site of persistence and a source for fecal shedding of certain enteric food-borne pathogens, such as Salmonella or Campylobacter spp. (3, 6). Stoffregen et al. (13) demonstrated that when calves were immunosuppressed and experimentally inoculated, E. coli O157:H7 localized in the gallbladder. They speculated that the gallbladder may be a site and source of gastrointestinal E. coli O157:H7 and contamination of meat at slaughter. Recently, Jeong et al. (10) reported isolation of E. coli O157:H7 in gallbladders of cattle and suggested that the organism can reside at a low level in gallbladders of cattle. The purpose of this study was to determine whether the gallbladders of cattle harbor E. coli O157:H7.The cattle sampled in this study were delivered to two commercial abbatoirs in the Midwest for slaughter. A total of 933 cattle were sampled on 11 different dates in the summer of 2005 (Table 1). Gallbladders were cut from the liver after evisceration of the carcass. The gallbladders were cut open, and bile was allowed to drain out. The tissue was inverted over a gloved hand of the collector, rinsed with tap water until free of visible bile, and then swabbed vigorously with a foam-tipped applicator (VWR International, Buffalo Grove, IL). The swab was then placed into a test tube containing 3 ml of gramnegative broth (BD, Franklin Lakes, NJ) containing cefixime, cefsulodin, and vancomycin (GNccv) and placed on ice (9). The entire gallbladder was then placed into a Whirl-Pak bag (Nasco, Ft. Atkinson, WI), and the bags were immediately placed on ice. Feces were accessed by making a full-thickness, longitudinal incision through the distal ...

TCEC is a successful, simple treatment for canine distichiasis.

A 2-year-old male castrated Domestic Short-haired cat presented to the Ophthalmology Service at the Matthew J. Ryan Veterinary Hospital at the University of Pennsylvania for evaluation of chronic bilateral ocular discharge and blepharospasm. Initial ophthalmic examination revealed severe conjunctivitis and keratitis and the presence of upper eyelid distichiae bilaterally. Initial therapy for suspected feline herpesviral infection provided moderate, but not complete, resolution of the clinical signs. Over the subsequent year, the cat suffered from recurrent, severe, ulcerative keratitis in both eyes despite appropriate medical therapy. Approximately 13 months after the initial presentation, the distichiae were surgically removed using transconjunctival electrocautery, which resulted in complete resolution of the clinical signs. This report documents bilateral distichiasis in a cat, a condition that is considered rare in this species.