Progesterone inhibits the in vitro L3/L4 molting process in Haemonchus contortus

We evaluated the direct effects of progesterone on the morphology, maturation and behavior of Haemonchus contortus larvae in vitro. The presence and location of possible progesterone receptors in these larvae were also determined. The addition of 8ng/mL of progesterone to larval cultures over 10days reduced larval enlargement, while the addition of 160ng/mL of the hormone increased the enlargement. Up to 62% and 65% of the H. contortus larvae molted from third-stage larvae (L3) to fourth-stage larvae (L4) when cultured in RPMI-1640 media without hormone for 5 and 10days, respectively. The addition of different progesterone concentrations (1, 8, 16, 80 and 160ng/mL) to the larval cultures significantly inhibited the molting process within the same periods. The addition of 8ng/mL or higher progesterone concentrations to the cultures significantly increased larval motility (p<0.05) compared with unstimulated larvae. Flow cytometry showed the expression of progesterone receptors (P4-R) in 15% of the cells from newly isolated H. contortus larvae. When the larvae were cultured for 5days in the presence of the hormone, the percentage of P₄-R+ cells remained the same. In contrast, unstimulated larvae showed a significant reduction in the number of P₄-R+ cells. Using confocal microscopy, a greater concentration of P₄-Rs was immunolocated in the anterior portion of the alimentary tract of the larvae, suggesting that the cells in this region are targeted by the hormone. The results of the present study show that H. contortus larvae have possible P₄-Rs and respond to this hormone by inhibiting their molting process, thereby suggesting the participation of progesterone in the larval arrest phenomenon.