Use of may-grunwald giemsa stain for microscopic evaluation of cell culture inoculated with microsporidia in vitro

Pereira, A.1,2, Saraiva, A.M.A1 and Lallo, M.A.1,2

Use of may-grunwald giemsa stain for microscopic evaluation of cell culture inoculated with microsporidia in vitro

Pereira, A.1,2, Saraiva, A.M.A1 and Lallo, M.A.1,2

1Paulista University 2Centro UniversitÃƒÂ¡rio SÃƒÂ£o Camilo, Brazil

: J Immunol Tech Infect Dis

Abstract

Microsporidia are obligatory intracellular parasites that infect human and other animals. These pathogens have been recognized as opportunistic parasites of immunodeficient patients and after the advent of HIV and AIDS, interest in the in vitro culture of them has been increasingly. In our laboratory we have been used in vitro culture to study the species Encephalitozoon cuniculi and E. intestinalis in monkey and rabbit kidney cell lines (Vero and RK-13). Recently we investigated the May-Grunwald Giemsa stain for microscopic evaluation of cells inoculated with E. cuniculi in vitro. Cell lines were incubated in sterile circular glass coverships in 24-well plates and were inoculated with spores of E. cuniculi. After the inoculation and incubation, the coverslips were fixed with methanol, stained with May-Grunwald Giemsa, mounted on glass slides and examined with a light microscope. Each entire coverslip was scanned at a magnification of X 1000. May-Grunwald Giemsa stain permitted the visualization of parasitophorous vacuoles (PV) containing mature microsporidian spores in the cytoplasm of the infected cells. The stain used in this study are commonly used in hematology laboratory routine but has not been used to identify microsporidia spores. This result suggests that May-Grunwald Giemsa is adequate for visualize the PV containing spores of microsporidia inside the infected cell cultures to study this pathogen in vitro.