一种大黄鱼盾纤毛虫病PCR检测方法的建立与应用OA

In order to prevent outbreaks of scuticociliatosis of juvenile large yellow croaker(Larimichthys crocea)and ensure that parasite-free juveniles are stocked in sea cages,species-specific primers targeting the 18S rRNA gene sequences of Miamiensis avidus and Uronema marinum were designed to establish a multiplex PCR detection method for scuticociliatosis.The method was comprehensively evaluated through primer specific-ity verification,sensitivity testing,and application to naturally infected samples.The results demonstrated that the method could specifically amplify DNA from M.avidus and U.marinum,without cross-reactivity with non-target pathogens such as Trypanosoma,Pseudomonas plecoglossicida,and Iridovirus,nor with DNA from healthy L.crocea tissues,indicating good specificity.Sensitivity tests revealed that the minimum detectable concentrations for M.avidus and U.marinum were 4.82×103 copies/μL(2.4 ng/L)and 1.67×103 copies/μL(1.3 ng/L),re-spectively,demonstrating high sensitivity.Ten naturally infected croaker samples were tested using the PCR meth-od,and six positive samples were detected in total,toutperforming traditional microscopy which detected only five,hus further confirming its diagnostic accuracy and sensitivity in field samples.