Environmental effects on sex determination in Litopenaeus vannamei were studied by rearing day 1 postlarvae at three temperatures, under three photoperiods, at high density and by starving. None of the environmental conditions affected sex determination or differential development of gender in this species. From day 50, the development of the endopodite of the first pair of pleopods revealed the first external differentiation, showing a triangular structure with three setae in females, whereas a tubular structure remained in males. Juvenile shrimp sex differentiation took place from days 50–90, independent of size, only if postlarvae reached a development threshold of 150 mg of body weight and 20 mm of body length previously. Histology and scanning electron microscopy of the vas deferens revealed that the androgenic gland (AG) is a single 2-mm cord attached in the subterminal ejaculatory region, just before the distal vas deferens narrows. The AG is composed of large oval cells containing vacuolated cytoplasm, and each cell has a prominent rounded nucleus, similar to all descriptions of the AG in Malacostracans, so we assume that it should have the same function in sex differentiation.