[目的]建立能同时鉴测猪流感病毒（SIV）、猪乙型脑炎病毒（JEV）、猪瘟病毒（CSFV）、猪繁殖与呼吸综合征病毒（PRRSV）的多重荧光RT-PCR方法。[方法]根据4种病原体的高度保守基因序列，设计相应引物和探针，并对引物和探针浓度进行优化，检测48例疑似样本。[结果]建立了多重荧光RT-PCR方法，检测48例临床样本中，4种病毒均未感染的1例，SIV与JEV混合感染占2.1%，CSFV与PRRSV混合感染为31.3%。[结论]多重荧光RT-PCR对四种病原体的检测准确性好，具有快速的特点。%Objective] To develop a multiplex fluorescent RT-PCR assay to detect swine influenza virus(SIV), Japanese encephalitis virus(JEV),classical swine fever virus(CSFV),and porcine reproductive and respiratory syndrome virus(PRRSV). [Method] Specific primers and probe were designed based on the highly conserved gene sequences of the four swine pathogens and their working concentrations and the amplification conditions were opti-mized. The multiplex fluorescent RT-PCR assay was evaluated with 48 suspected clinical samples. [Result] The multiple fluorescent RT-PCR assay was established for 4 swine viruses. Only one out of 48 samples was negative to the four viruses;the positive rate of SIV and JEV mixed infection was 2.1 %;the positive rate of CSFV and PRRSV mixed infection was 31.3 %.[Conclusion] The development of the multiplex fluorescent RT-PCR provided a sensitive,specific and rapid method to detect SIV,CSFV,PRRSV and JEV.
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