Ostrowski M, Mundo S L, Harris N B, Barletta R G, Lopez O J
School of Veterinary Sciences, University of Buenos Aires, Buenos Aires, Argentina.
Scand J Immunol. 2003 Nov;58(5):511-21. doi: 10.1046/j.1365-3083.2003.01334.x.
Mycobacterium avium ssp. paratuberculosis (M. paratuberculosis) causes Johne's disease, a chronic and fatal enteritis in ruminants. In the last stage of the disease, antibody titres rise and levels of interferon-gamma decrease, suggesting that the host-immune response is switching from a T helper 1 (Th1) to a Th2 profile. In infected cattle, the membrane protein p34 elicits the predominant humoral response against M. paratuberculosis. To map the B-cell epitopes of this antigen, affinity-purified bovine antibodies against the carboxy-terminal region of p34 were used to screen a 12-mer phage display library. Several phage clones carrying peptides resembling fragments of p34 were affinity selected. Based on the predicted amino acid sequence, peptides were chemically synthesized, which demonstrated reactivity with serum from naturally infected and p34-vaccinated cattle. Immunization of mice with these peptides elicited an anti-p34 antibody response. Two B-cell epitopes were identified and characterized. Based on the reactivity and the type of immune response elicited, epitope A was determined to be conformational, whereas epitope B was demonstrated to be sequential. Both epitopes were shown to be present in p34 proteins from M. avium ssp. avium or M. paratuberculosis but absent from M. intracellulare, the other member of the M. avium complex. Furthermore, both epitopes were mapped to regions of p34 that display high variability when compared to homologous proteins from other mycobacterial species of public and animal health importance. We hypothesize that these variable regions of p34 may play a role in the immunobiology of M. paratuberculosis infections.
副结核分枝杆菌（M. paratuberculosis）可引发反刍动物的慢性致命性肠炎——副结核病。在疾病的最后阶段，抗体滴度升高而γ干扰素水平降低，这表明宿主免疫反应正从辅助性T细胞1（Th1）型向Th2型转变。在受感染的牛中，膜蛋白p34引发针对副结核分枝杆菌的主要体液免疫反应。为了绘制该抗原的B细胞表位图谱，使用针对p34羧基末端区域的亲和纯化牛抗体筛选一个12肽噬菌体展示文库。筛选出了几个携带类似于p34片段的肽的噬菌体克隆。根据预测的氨基酸序列，化学合成了肽，这些肽与自然感染和接种p34疫苗的牛的血清具有反应性。用这些肽免疫小鼠引发了抗p34抗体反应。鉴定并表征了两个B细胞表位。根据引发的反应性和免疫反应类型，确定表位A为构象表位，而表位B为线性表位。已证明这两个表位存在于鸟分枝杆菌亚种鸟分枝杆菌或副结核分枝杆菌的p34蛋白中，但在鸟分枝杆菌复合群的另一个成员胞内分枝杆菌中不存在。此外，与其他对公共卫生和动物健康具有重要意义的分枝杆菌物种的同源蛋白相比，这两个表位均定位于p34中显示出高变异性的区域。我们推测，p34的这些可变区域可能在副结核分枝杆菌感染的免疫生物学中发挥作用。
