Location of polysaccharide on Chlamydia psittaci by silver-methenamine staining and electron microscopy

Previous serological studies have indicated that the group antigen of chlamydial organisms is composed of an acidic polysaccharide and a lipid component. The present study was undertaken in an effort to locate this polysaccharide complex by use of electron microscopy and a silver-methenamine marker. The meningopneumonitis strain of Chlamydia psittaci was propagated in HeLa-M cell culture. Organisms were purified by differential centrifugation, treatment with Genetron, and by gel filtration. After fixation and embedding, sections were obtained for electron microscopy. Sections were stained for carbohydrates with silver-methenamine. A double layer of regularly spaced silver grains of uniform size was observed at the periphery of the sectioned organisms tracing the contours of the surface membrane (cell wall). This intensity of staining was observed only when sections were oxidized with periodate prior to silver-methenamine staining. Prior treatment with 1% sodium deoxycholate resulted in a significant reduction in staining. It is considered probable that the periodate-sensitive polysaccharide found at the periphery of the organisms represents, or is a component of, the group antigen of these organisms.