Study On The Method Of A Latex Agglutination Test Detecting Antibodies Against Pasteurella Multocida Toxin

Swine infectious atrophic rhinitis(AR) is a widely epidemic,highly contagious, and severe disease of the upper respiratory tract of swine, which Tesults in bad turbinate atrophy and degeration, reduced daily weight gain, inefficient feed conversion and increased time to market and applys to other clinical infections like PRRS. It is known that toxingenic Pasteurella multocida (T~+Pm)and I Bordetella bronchiseptica (Bb)are considered as the important primary pathogens;moreover, Pasteurella multocida toxin(PMT) including the most noxtious dermonecrotic toxin (DNT) encoded by toxin A, in particular, is the major nosogenetic factor. With the intensivism breeding of swine production speeding up, AR leads to a great loss for swine industry. So, the establishment of a rapid and special seradiagnosis of detecting antibodies against Pasteurella multocida toxin will pave a scintific basis for its prevention and cure.A method of latex agglutination test rapidly detecting antibodies to Pasteurella multocida toxin (PMT)was established with C terminal protein of Pasteurella multocida toxin which was prokaryoticly expressed by the plasmid of pET28bC2115 transformed into E.coli BL21.The contents of studies as follows:1.Transformed into E.coli BL21, the plasmid of pET28bC2115 obtained efficient expression as nucleocapsid protein in E.coli BL21 with an approximate 78kDa protein which had biological activity identified by Western-blot analysis.2.Conjugaing fine concentration of the purified protein from pET28bC2115 to carboxylate-modified microspheres via carbodiimide(EDC)-mediated coupling reactions, the diagnostic antigen of PMT was obtained and a latex agglutination test(LAT) for rapidly detecting antibody to PMT was developed with its specificity and stability studied. The experiment that the diagnostic antigen was preserved in 4℃ and room temperature(RT) respectively denoted that it could be stored for at east 6 months in 4℃ and 3 months in RT and was stable.3.The developed LAT was compared with ELISA screened antibodies to PMT by simultaneously detecting 83 pig serum samples from 9 farms. The coincidence rate of both assays is 94.0%(78/83) without significant difference between them and therefore the LAT of testing antibodies to PMT was established.4.Meanwhile 1680 pig sera from such provinces as Guangzhou, Fujian, Shanghai, Hunan, Hubei and Shangdong was tested by the LAT; its total positive rate is 28.3% and hitted 33.7% in Januaray, mainly from sow and piglets. The results showed excellentagreement with AR epidemiologic trait.The above study proves that the LAT of T^Pm is special and reduplicating by the comparision with the ELISA. The latex antigen of PMT is stable and is suitable for storage and conveyance by the preserved experiment.lt is revealed that the developed LAT is of high speciality and simplicity and makes the basis of serological diagnosis of PMT,so as to eradicate it finally.