猪肺炎支原体气溶胶富集检测技术的初步研究

The Preliminary Study of Collection and Detection Technology of Mycoplasma hyopneumoniae in Aerosol

为了建立一个简单易行的猪肺炎支原体气溶胶富集和检测技术,结合液体冲击式采样器和过滤式采样器的原理,自行设计一个气溶胶双重富集装置,并在密闭环境下采集人工制备的猪肺炎支原体气溶胶,荧光定量PCR检测以确定该装置的采集效率。再通过收集不同猪群中的气溶胶进行检测以确定该装置临床应用的可行性。首先,收集人工感染Mhp阴性仔猪后不同时间猪舍内的气溶胶;其次,收集1个猪支原体肺炎发病猪场和1个阳性未发病猪场的产房、保育舍和育肥舍共11个猪舍的气溶胶,荧光定量PCR或套式PCR检测所收集的气溶胶。结果显示:实验室模拟采样的采集效率达到(37.04±6.43)%,人工感染猪肺炎支原体后7天即在饲养房间的气溶胶中检到猪肺炎支原体;2个猪场共11个不同猪舍气溶胶中猪肺炎支原体检测结果均为阳性,检出率100%。成功建立了一个简单易行的猪场猪肺炎支原体气溶胶富集及检测技术。

The research was to establish a simple method for collecting and detecting Mycoplasma hyopneumoniae(Mhp) in aerosol.Based on the mechanism of liquid impact type sampler and filtration sampler,an double concentration aerosol sampler was design for collecting the aerosol of Mycoplasma hyopneumoniae.Firstly,the collection was performed in a closed environment full of artificial aerosol of Mhp.Secondly,collection efficiency was detected by Real-time PCR method.Thereafter,the clinical feasibility of the designed equipment was tested by collecting aerosol samples in different pig herds.In one assay,the samples were collected at different times from one pig house challenged with Mhp.In another assay,the samples was collected from the delivery room,nursery and fattenning house of one MPS outbreak farm as well as a Mycoplasma hyopneumoniae infection positive pig farm without obvious clinical.All the aerosol samples were then detected by real-time PCR or nest PCR.The results showed that,the collection efficiency of the designed bioaerosol sampler was(37.04±6.43) %,Mhp could be detected 7 days after intratracheal challenge with pneumonic lung homogenate suspension.Aerosol samples of 11 pig houses from the two Mhp positive pig farm with or without clinical symptoms all showed a positive result of PCR,the positivity rate was 100%.In conclusion,one simple and easy method for collecting and detecting Mycoplasma hyopneumoniae in aerosol was successfully founded which was practical for clinical use.