Sex and culture duration differences in Ascaridia galli Extracellular Vesicles

To determine how to obtain the highest amount of EVs shed by A. galli, several kinetic experiments were conducted. Cleaned and sexed worms were grouped according to size. For both females and males, medium-sized worms (females: 5.7-6.8 cm; males: 3.7-4.8 cm) produced the most particles per mL, indicating that EV production depends on life stage.
The amount of EVs shed by A. galli was relatively stable across replicates, though the optimal culture duration for maximal EV yield differed between sexes. Female worms produced the highest average particle yield after 24 hours of culture, with no significant difference compared to 48 hours. Male worms shed the highest number of EVs after 40 hours, but again, no significant difference was found between timepoints.
No morphological differences were observed between EV fractions from different timepoints or worm sexes. Following size-exclusion chromatography, EVs from females were mainly detected in fractions 5-8, with the highest yield in fraction 6. In contrast, male worms produced significantly fewer EVs overall, and none were detected in fraction 8.
The size distribution of EVs did not differ between sexes, with most particles ranging from 100 to 200 nm.