CRYPTOCOCCAL CELL ENLARGEMENT IN MURINE LUNG TISSUE
Document Type
Presentation
Start Date
22-10-2010 3:00 PM
End Date
22-10-2010 4:30 PM
Description
Cryptococcus neoformans is a human fungal pathogen which enters via the lungs and may disseminate to cause meningoencephalitis in immunocompromised individuals. Two _g_ryptococcal regulators of g-protein signaling (Crg) have been identified, and deletion of CRG2 decreases virulence in a mouse model of infection. Recent studies have also identified a novel cell morphology during pulmonary infection which is linked to virulence, and preliminary examination of crg2 in lung tissues of infected mice revealed a potential defect in enlarged cell formation. Under in vitro conditions, cryptococcal cells reach a diameter of 2 to 5 uM which may increase up to 10 fold in pulmonary tissues during infection. To test whether deletion of CRG2 affected the ability of C. neoformans to enlarge within the lung to greater than 15 uM diameter, mice were inoculated with equivalent suspensions of wild type (WT) or crg2 cells and observed for 2, 7, and 14 days post infection, or until moribund. Lung tissues were collected for colony forming unit (CFU) determination or formalin fixation at each time point. Lungs were weighed, homogenized and diluted serially for CFU determination. Formalin fixed lungs were processed, paraffin embedded, sectioned at 5 uM, adhered to glass slides and stained with Periodic Acid Schiff's reagent (PAS) by standard means. Slides were examined in a blinded manner and five random fields photographed. PAS stained cryptococcal cells were measured in Fiji for area and Feret's diameter. CFU for crg2 infected animals were significantly decreased as compared to WT at all times assayed and, correspondingly, fewer cells were visualized in histological sections at all times assayed as well. Median diameter and area were not significantly different between strains, though both diameter and area did increase significantly from one time point to another for WT infected animals, a trend not seen in crg2 infected samples. Median Feret's diameter increased at 7 and 14 days post infection for WT and crg2, indicating a shift to enlarged cell morphology which was not evident in tissues from moribund animals. Overall, our evidence does not support a role for Crg2 in the formation of enlarged cryptococcal cells during pulmonary infection.
Recommended Citation
Whittington, Amy and Wang, P., "CRYPTOCOCCAL CELL ENLARGEMENT IN MURINE LUNG TISSUE" (2010). Dr. Joseph M. Moerschbaecher, III Graduate Research Day. 26.
https://digitalscholar.lsuhsc.edu/grad_rs/2010/poster2/26
CRYPTOCOCCAL CELL ENLARGEMENT IN MURINE LUNG TISSUE
Cryptococcus neoformans is a human fungal pathogen which enters via the lungs and may disseminate to cause meningoencephalitis in immunocompromised individuals. Two _g_ryptococcal regulators of g-protein signaling (Crg) have been identified, and deletion of CRG2 decreases virulence in a mouse model of infection. Recent studies have also identified a novel cell morphology during pulmonary infection which is linked to virulence, and preliminary examination of crg2 in lung tissues of infected mice revealed a potential defect in enlarged cell formation. Under in vitro conditions, cryptococcal cells reach a diameter of 2 to 5 uM which may increase up to 10 fold in pulmonary tissues during infection. To test whether deletion of CRG2 affected the ability of C. neoformans to enlarge within the lung to greater than 15 uM diameter, mice were inoculated with equivalent suspensions of wild type (WT) or crg2 cells and observed for 2, 7, and 14 days post infection, or until moribund. Lung tissues were collected for colony forming unit (CFU) determination or formalin fixation at each time point. Lungs were weighed, homogenized and diluted serially for CFU determination. Formalin fixed lungs were processed, paraffin embedded, sectioned at 5 uM, adhered to glass slides and stained with Periodic Acid Schiff's reagent (PAS) by standard means. Slides were examined in a blinded manner and five random fields photographed. PAS stained cryptococcal cells were measured in Fiji for area and Feret's diameter. CFU for crg2 infected animals were significantly decreased as compared to WT at all times assayed and, correspondingly, fewer cells were visualized in histological sections at all times assayed as well. Median diameter and area were not significantly different between strains, though both diameter and area did increase significantly from one time point to another for WT infected animals, a trend not seen in crg2 infected samples. Median Feret's diameter increased at 7 and 14 days post infection for WT and crg2, indicating a shift to enlarged cell morphology which was not evident in tissues from moribund animals. Overall, our evidence does not support a role for Crg2 in the formation of enlarged cryptococcal cells during pulmonary infection.
Comments
See abstract book page 70