Four isolates of infectious haematopoietic neerosis virus (IHNV) were obtained from rainbow trout, Oncorhynchus mykiss (Walbaum), and masu salmon, Oncorhynchus mason (Walbaum), fry during epizootics at hatcheries in Korea. The four isolates of IHNV were compared with three from salmonids in the USA (SRCV, OSV and RB-76) by analysis of virion proteins in sodium dodecyi sulphate polyacrylamide gel electrophoresis (SDS-PAGE) and neutralization tests, with two monoelonal antibodies raised against SRCV (MAb SRCV/A4) and RB-76 (MAb RB/B5). Based on the antigenieity and the size of the structural proteins, one Korean isolate from masu salmon (SCS) is similar to RB-76 and is an electropherotype 1. The other three isolates from rainbow trout (PRT, YRT and MRT) were identical to eaeh other in the mobilities of their virion proteins in SDS-PAGE, and although their nueleocapsid (N) proteins comigrated with that of the RB-76 isolate, they differed from RB-76 in the smaller matrix 2 (M2) protein they contained. In addition, the three Korean isolates (PRT, YRT and MRT) could be divided into two groups by reactivity with MAb RB/B5. While the YRT isolate was neutralized by this MAb, the PRT and MRT isolates were not. suggesting that there are at least two neutralizing antigenic variants of IHNV in Korea.

MATERIALS AND METHODS Antigen preparationPorcine adrenocorticotropic hormone (ACTH) extracted from the pituitary (Sigma Chemical Co., Saint Louis, MO, USA) was conjugated to keyhole limpet hemocyanin (KLH) or human histone carrier using either 3-maleimidobenzoic acid N-hydroxysuccinimide (MBS) or glutaraldehyde, under a 2 × 2 × 2 factorial arrangement of treatments; main effects included hapten (ACTH vs none), carrier and crosslinker. The hapten-carrier conjugation using MBS was performed following the procedure of Liu et al. (1979). Briefly, 70 mg KLH or histone dissolved in 4.4 ml of 50 mM phosphate-buffered saline (PBS), pH 7.4, was mixed with 875 µl of 6.0 mg MBS/ml N, N-dimethyl formamide solution for 30 min on a rocker at room temperature. After removing free MBS on a P-6 disposable gel filtration column (Bio-Rad, Hercules, CA, USA), fractions corresponding to activated carrier were pooled and mixed with 13.2 ml PBS containing 70 mg dissolved ACTH or none for 3 h on a stirring plate. The hapten-carrier