Litopenaeus vannamei collected from Thailand (T) and the United States (M) were uesd as parents, four progeny populations of TT, MM, TM and MT were constructed by diallel cross, with a total of 20 families. The tolerance of young shrimp to high ammonia-N, high pH and low salt was compared through 96 h acute toxicity test, the heterosis of each mating combination was analyzed, and the tolerance of parents and offspring was evaluated.The results showed that under 96 h of high ammonia-N, high pH, and low salt stress, the mortality rates of each family were 19.52%–92.22%, 23.29%–92.58%, and 19.95%–80.17%, respectively. There were significant differences in the tolerance of different families to ammonia-N, pH, and salinity stress (P < 0.05). The population with a female parent from the United States has stronger tolerance to ammonia-N, pH, and low salt stress than the population with a female parent from Thailand. The population with a male parent from Thailand has weaker tolerance to pH and low salt stress than the population with a male parent from the United States, but is superior to the population with a male parent from the United States under ammonia-N stress. The heterosis rates of the hybrid population TM in acute high ammonia-N, high pH and low salinity were 81.67%, 44.89% and −10.18%, respectively; The heterosis rate of MT population was 14.89%, 38.82% and −8.09%, respectively. The overall resistance of the four populations showed MM > MT > TT > TM. The population TM has obvious heterosis in high ammonia-N and high pH tolerance traits, and the family MM7 has strong low salt tolerance, so it can be considered as a candidate family for subsequent breeding work. The experimental results provide a basis for screening new strains of vannamei shrimp with strong stress resistance through family breeding.