The infectious hypodermal and hematopoietic necrosis virus (IHHNV) is very pathogenic for Litopenaeus stylirostris whereas infection in Litopenaeus vannamei is known to induce development and growth abnormalities and cause economic losses that range between 10% and 50% (Lightner and Redman, 1998). In the present work, on the basis of nested-PCR analysis, IHHNV prevalences were determined to be between 47% and 63% in Ecuadorian wild and domesticated broodstocks and around 95% in Panamanian domesticated broodstock. IHHNV was regularly detected in the ovaries of infected females whereas sperm from infected males was generally free of virus. IHHNV vertical transmission from infected females was clearly established. In the case of highly infected females, embryo development may abort. IHHNV-free nauplii and larvae were produced from females and males that were found to be free of virus on the basis of nested PCR performed in the case of females after eyestalk ablation and the first spawning. The reliability of this testing process was shown to be very high since about 87% of females was confirmed IHHNV-free through a second nested-PCR analysis performed after some additional spawnings. The nauplius productivity was higher for IHHNV-free females than for the infected ones. Virus-free verification of L. vannamei broodstock may become a general routine hatchery procedure in the near future to prevent the disease from spreading through vertical transmission. © 2003 Elsevier Science B.V. All rights reserved.