A lipoprotein-cholesterol-albumin serum substitute stimulates Giardia lamblia encystation vesicle formation

We found previously that the A6 clone of Giardia lamblia strain WB that did not encyst in vitro was blocked at an early stage in differentiation, as it did not form encystation secretory vesicles (ESV) efficiently or express cyst antigens, in comparison with the related clone C6. We now report that A6 formed ESV normally in the suckling mouse model. Therefore, we asked whether our serum-containing encystation media might lack a stimulus or component or contain an inhibitor of ESV formation to which A6 was especially sensitive. We found that replacing bovine serum with a lipoprotein-cholesterol solution and bovine serum albumin (LPC) in pre-encystation and encystation media increased ESV formation by both A6 and C6. The % of A6 cells with ESV increased from 8% in BS medium to 48% in LPC medium, compared with 64% and 98% for C6. Similarly, the average number of ESV/positive cell increased from 1.5 in BS medium to 7.7 in LPC medium for A6, and from 13.3 to 19.7 for C6. Moreover, in LPC encystation media, A6 expressed the cyst wall epitope recognized by monoclonal GCSA-1. Although formation of water-resistant cysts by A6 was increased > 60 fold in LPC media, the numbers of cysts remained only approximately 3-15% that of C6. This suggests that LPC may primarily affect early events in encystation and that A6 may require additional factors later in encystation.