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Justine R. Smith, Kathleen Mohs, Liam M. Ashander, Joao M. Furtado; Imaging Murine Ocular Toxoplasmosis with Topical Endoscopic Fundus Imaging (TEFI). Invest. Ophthalmol. Vis. Sci. 2011;52(14):2937.
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Ocular toxoplasmosis is a common retinal infection affecting up to 15% of the population in high-risk areas of the world. We investigated the impact of Toxoplasma gondii strain on murine ocular toxoplasmosis, including a clonal isolate (RH), and natural isolates of common haplogroups 1 (GT-1) and 2 (DEG) and exotic haplogroup 6 (TgCatBR2 and GPHT), using TEFI for the first time to visualize the disease in vivo.
Female C57BL/6 mice aged 8 - 12 weeks were injected intravitreally OD with T. gondii tachyzoites (104/2 µL saline) or saline alone (n=5-7 mice/group) according to a protocol modified from Charles et al (Infect Immun, 2007). Six days post-injection, fundus imaging was performed under general anesthesia, with dilated pupils, using TEFI, per Paques et al (Invest Ophthalmol Vis Sci, 2007) and Xu et al (Exp Eye Res, 2008) with: tripod-mounted Nikon D90 camera in manual focus mode; 60 mm F/2.8 D lens; 58-52 step-down ring; Storz 481C halogen light; and Storz 1218 tele-otoscope. Five images per eye, including posterior pole, and superior, nasal, temporal and inferior peripheries, were obtained. Photographs were saved in RAW mode and formatted with Nikon ViewNX2 software. Image montages were scored using numerical grading that considered status of vitreous, retina and retinal vasculature, with maximum severity score of 12. Post-mortem, eyes were enucleated, formalin-fixed, paraffin-embedded, and sectioned at 5µm. Sections were stained with hematoxylin and eosin.
Intraocular inflammation in infected mice was characterized clinically by white lesions with fluffy borders, retinal vasculitis and/or vitreous haze. Some parasite- and saline-injected mice had atrophic retinal lesions, possibly related to raised intraocular pressure at the time of injection, that were not considered in scoring. Inflammation was significantly different between saline controls and infected mice for all parasite strains (p≤0.02), and varied across infected groups, with mean severity scores of: RH, 5.0; GT-1, 3.8; DEG, 2.0; TgCatBR2, 4.6; and GPHT, 5.0. Histopathology demonstrated a mononuclear infiltrate involving vitreous, retina and/or optic nerve, with severity matching clinical findings.
TEFI is a new intravital imaging modality that we show to be a useful method for evaluating murine ocular toxoplasmosis; clinical findings are consistent with histopathological observations. Our results, comparing inflammation in different parasite strains, suggest that parasite genetics influences the severity of this disease.
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