In vitro expression of the 50-kDa and 30-kDa fragments of the P97 adhesin of Mycoplasma hyopneumoniae in Escherichia coli and their use for serodiagnosis.
This article reports the cloning and expression of 2 fragments of the P97 adhesin of Mycoplasma hyopneumoniae for use in serodiagnosis: a 50-kDa fragment (including the N-terminal cleavage site) and a 30-kDa fragment (including the C-terminal R1 and R2 repeats, which are essential for adherence). The genes encoding the fragments were amplified, cloned, and expressed in the Escherichia coli expression system BL21 (DE3)pLysS. Antiserum against the purified recombinant proteins reacted with the mycoplasmal 97-kDa intact protein and the 66-kDa major cleavage fragment, confirming that both cloned fragments could induce antigen-specific antibodies in mice. Of 70 serum samples from nonvaccinated pigs, 26 (37%) were seropositive when the 30-kDa fragment was used as an antigen for enzyme-linked immunosorbent assay, suggesting that natural mycoplasmal infection is quite common in Korea. However, only 4 samples were seropositive when the 50-kDa fragment was used; this fragment was therefore deemed unsuitable for serodiagnosis. The 30-kDa fragment protein might be useful for measuring antibody response to vaccination and for detecting mycoplasmal infection.
Jang
Eun Jin
EJ
College of Veterinary Medicine, Chonnam National University, Gwangju 500-757, Republic of Korea.
Kim
Tae Jung
TJ
D018829:Adhesins, Bacterial / Q000235:genetics*; D000818:Animals; D000907:Antibodies, Bacterial / Q000097:blood / Q000276:immunology; D000942:Antigens, Bacterial / Q000276:immunology; D001422:Bacterial Adhesion / Q000502:physiology*; D004926:Escherichia coli; D008970:Molecular Weight; D045705:Mycoplasma hyopneumoniae / Q000502:physiology*; D045729:Pneumonia of Swine, Mycoplasmal / Q000175:diagnosis*; D016133:Polymerase Chain Reaction / Q000379:methods / Q000662:veterinary; D012698:Serologic Tests / Q000379:methods / Q000662:veterinary; D013552:Swine
D018829:Adhesins, Bacterial; D000907:Antibodies, Bacterial; D000942:Antigens, Bacterial; C092236:P97 protein, Mycoplasma