在采采蠅中腸中發(fā)現(xiàn)的前循環(huán)型布氏錐蟲的掃瞄電子顯微圖片,，圖片來自維基共享資源,。
布氏錐蟲(Trypanosoma brucei)導(dǎo)致的昏睡病(sleeping sickness)可通過采采蠅(tsetse fly)的叮咬傳染給人類(和動(dòng)物)。根據(jù)2012年2月15日發(fā)表在生物醫(yī)學(xué)中心(BioMed Central)旗下開放存取的Microbial Cell Factories期刊上的一篇新研究論文,，研究人員利用一種自然條件下生活在采采蠅體內(nèi)的一種細(xì)菌釋放納米抗體(nanobody,，一類抗體片段)對(duì)抗這種錐蟲。這就意味這種細(xì)菌在采采蠅體內(nèi)發(fā)揮著一種類似特洛伊木馬的作用,。這些結(jié)合到這種寄生蟲表面的納米抗體是制造能夠殺死或阻斷錐蟲發(fā)育的特定納米抗體的第一步,。
昏睡病威脅著生活在非洲撒哈拉沙漠以南地區(qū)上百萬人的生命。在布氏錐蟲感染第一階段(血淋巴階段),，它導(dǎo)致人發(fā)熱,、頭痛、關(guān)節(jié)疼痛和發(fā)癢,。當(dāng)這種寄生蟲跨越血腦屏障時(shí)便進(jìn)入感染第二階段(神經(jīng)階段),，它導(dǎo)致思緒混亂、較差的協(xié)調(diào)能力和睡眠紊亂,。若不進(jìn)行治療,，昏睡病是致命的。然而,，診斷和治療這種疾病比較困難,，只有接受過特別培訓(xùn)的人員才能開展。錐蟲感染牛后會(huì)導(dǎo)致牛貧血,，而且還能夠?qū)е滤鼈兯烙谪氀Y,。綜合在一起就是，這些已經(jīng)對(duì)整個(gè)非洲大陸的公共健康和農(nóng)業(yè)發(fā)展帶來嚴(yán)重影響,。
細(xì)菌(Sodalis glossinidius)是一種內(nèi)共生菌,，類似于定居于人腸道的“有益細(xì)菌”，存在于采采蠅的中腸,、肌肉,、脂肪和唾液腺之中。鑒于雌性采采蠅會(huì)把這些細(xì)菌傳給它的后代,，那么一旦將這種雌性蠅放到野外,，基因修飾的細(xì)菌也應(yīng)當(dāng)能夠在采采蠅之間代代相傳。比利時(shí)研究人員對(duì)這種內(nèi)共生菌進(jìn)行基因改良,，這樣它分泌一種單結(jié)構(gòu)域抗體結(jié)合到布氏錐蟲的一種可變表面糖蛋白(variant surface glycoprotein, VSG)上,。這種基因突變細(xì)菌的生長(zhǎng)不受影響，因此一旦放到大自然中會(huì)增加它們自己的存活率,。
比利時(shí)安特衛(wèi)普市熱帶疾病研究所的Van Den Abbeele教授解釋道,，“當(dāng)我們?cè)谀M采采蠅腸道內(nèi)部的條件下研究活著的錐蟲時(shí)，這種基因改造菌表達(dá)的納米抗體有生物活性，能夠結(jié)合到這種寄生蟲的整個(gè)表面上,。鑒于我們知道這種技術(shù)有效,，我們正在研究能夠在采采蠅腸道中破壞或阻斷這種寄生蟲發(fā)育的納米抗體。”
最近的流行性昏睡病發(fā)生于20世紀(jì)70年代,。盡管人們?cè)谶^去10年持續(xù)的努力和新增病例數(shù)量在下降，但是迄今為止這種疾病還沒有得到根除,。這種新技術(shù)為人們對(duì)抗這種破壞性的疾病提供希望,。(生物谷：towersimper編譯)
doi:10.1186/1475-2859-11-23
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Expression and extracellular release of a functional anti-trypanosome Nanobody(R) in Sodalis glossinidius, a bacterial symbiont of the tsetse fly
Linda De Vooght, Guy Caljon, Benoit Stijlemans, Patrick De Beatselier, Marc Coosemans and Jan Van Den Abbeele
Background
Sodalis glossinidius, a gram-negative bacterial endosymbiont of the tsetse fly, has been proposed as a potential in vivo drug delivery vehicle to control trypanosome parasite development in the fly, an approach known as paratransgenesis. Despite this interest of S. glossinidius as a paratransgenic platform organism in tsetse flies, few potential effector molecules have been identified so far and to date none of these molecules have been successfully expressed in this bacterium.
Results
In this study, S. glossinidius was transformed to express a single domain antibody, (Nanobody(R)) Nb_An33, that efficiently targets conserved cryptic epitopes of the variant surface glycoprotein (VSG) of the parasite Trypanosoma brucei. Next, we analyzed the capability of two predicted secretion signals to direct the extracellular delivery of significant levels of active Nb_An33. We show that the pelB leader peptide was successful in directing the export of fully functional Nb_An33 to the periplasm of S. glossinidius resulting in significant levels of extracellular release. Finally, S. glossinidius expressing pelBNb_An33 exhibited no significant reduction in terms of fitness, determined by in vitro growth kinetics, compared to the wild-type strain.
Conclusions
These data are the first demonstration of the expression and extracellular release of functional trypanosome-interfering Nanobodies(R) in S. glossinidius. Furthermore, Sodalis strains that efficiently released the effector protein were not affected in their growth, suggesting that they may be competitive with endogenous microbiota in the midgut environment of the tsetse fly. Collectively, these data reinforce the notion for the potential of S. glossinidius to be developed into a paratransgenic platform organism.
