Choi Changsun, Chae Chanhee
Department of Veterinary Pathology, College of Veterinary Medicine and School of Agricultural Biotechnology, Seoul National University, Kwanak-Gu 151-742, Seoul, Republic of Korea.
J Vet Diagn Invest. 2003 Jan;15(1):35-41. doi: 10.1177/104063870301500108.
A seminested reverse transcription-polymerase chain reaction (RT-PCR) was developed for the detection of classical swine fever virus (CSFV) in semen. Five boars were inoculated intranasally with CSFV isolate propagated in PK15 cells. Two boars inoculated with the supernatant of noninfected PK15 cells were kept as controls. Semen and serum samples were collected twice weekly for 63 days postinoculation (dpi). Samples were tested for the presence of antibodies to CSFV by an enzyme-linked immunosorbent assay and for the presence of CSFV nucleic acid by seminested RT-PCR. Antibodies to CSFV could be detected as early as 7 dpi in 1 boar, and all 5 infected boars were found positive by 14 dpi. CSFV from boar semen was infrequently identified by virus isolation compared with seminested RT-PCR. CSFV nucleic acid was detected in semen by seminested RT-PCR as early as 7 dpi in 3 infected boars and persistently thereafter in all 5 infected boars until 63 dpi. When separated fractions of CSFV-contaminated semen were analyzed by the seminested RT-PCR, the CSFV nucleic acid was detected mainly in seminal fluid and occasionally in nonsperm cells. CSFV antigen was also detected in nonsperm cells from semen smear by immunohistochemistry. Thus, infection via semen, specially through CSFV-infected seminal fluid, seems to be a major route of transmission of CSFV.
开发了一种半巢式逆转录聚合酶链反应（RT-PCR）用于检测精液中的经典猪瘟病毒（CSFV）。5头公猪经鼻接种在PK15细胞中传代的CSFV分离株。将2头接种未感染PK15细胞上清液的公猪作为对照。接种后63天内每周采集两次精液和血清样本。通过酶联免疫吸附试验检测样本中CSFV抗体的存在情况，通过半巢式RT-PCR检测CSFV核酸的存在情况。在1头公猪中最早于接种后7天可检测到CSFV抗体，到接种后14天所有5头感染公猪均呈阳性。与半巢式RT-PCR相比，通过病毒分离很少能从公猪精液中鉴定出CSFV。通过半巢式RT-PCR最早在3头感染公猪的精液中于接种后7天检测到CSFV核酸，此后在所有5头感染公猪中持续存在直至接种后63天。当用半巢式RT-PCR分析CSFV污染精液的分离组分时，CSFV核酸主要在精液中检测到，偶尔在非精子细胞中检测到。通过免疫组织化学在精液涂片的非精子细胞中也检测到了CSFV抗原。因此，经精液感染，特别是通过感染CSFV的精液感染似乎是CSFV的主要传播途径。
