Intact Giardia muris
cysts were subjected to consecutive
chloroform/
methanol and 2%
sodium dodecyl sulfate (SDS) extractions, and to
amyloglucosidase treatment. The SDS-insoluble,
amyloglucosidase-fast
cyst walls (ACW) were further incubated with
chymotrypsin, trypsin,
papain, or
pronase. Low voltage scanning electron microscopy revealed no discernible change in the ultrastructure of the filamentous layer of the
cyst wall following any of these treatments. Affinity for
cyst wall-specific
monoclonal antibody (Meridian Diagnostics, Cincinnati,
OH) was also retained after all treatments.
Periodic acid-Schiff staining and gas chromatography/mass spectrometry (GC/MS) of intact and treated
cyst hydrolysates showed a significant reduction in the amount of
glucose associated with the
cyst (72 nmoles/10(6) intact
cysts vs 1.9 nmoles/10(6) ACW) as a result of
amyloglucosidase treatment, indicating that
glucose is stored within Giardia as an SDS-insoluble
polymer.
Galactosamine was identified by GC/MS as the predominant
sugar associated with both the ACW and the
proteinase treated ACW (42 nmoles/10(6) ACW). High performance liquid chromatographic analysis of
amino acids from intact and treated
cyst hydrolysates revealed a marked reduction, but not elimination, of detectable quantities of identifiable
amino acid residues (255 nmoles/10(6) intact
cysts vs 6.8 nmoles/10(6)
proteinase treated ACW). These results suggest that the filamentous layer of the
cyst wall is primarily a
carbohydrate peptide complex.