Australian Animal Health Laboratories, CSIRO Livestock Industries, Geelong, Victoria, Australia.
PLoS One. 2011 Jan 25;6(1):e14576. doi: 10.1371/journal.pone.0014576.
The cane toad, Bufo (Chaunus) marinus, is one of the most notorious vertebrate pests introduced into Australia over the last 200 years and, so far, efforts to identify a naturally occurring B. marinus-specific pathogen for use as a biological control agent have been unsuccessful. We explored an alternative approach that entailed genetically modifying a pathogen with broad host specificity so that it no longer caused disease, but carried a gene to disrupt the cane toad life cycle in a species specific manner.
METHODOLOGY/PRINCIPAL FINDINGS: The adult beta globin gene was selected as the model gene for proof of concept of autoimmunity as a biocontrol method for cane toads. A previous report showed injection of bullfrog tadpoles with adult beta globin resulted in an alteration in the form of beta globin expressed in metamorphs as well as reduced survival. In B. marinus we established for the first time that the switch from tadpole to adult globin exists. The effect of injecting B. marinus tadpoles with purified recombinant adult globin protein was then assessed using behavioural (swim speed in tadpoles and jump length in metamorphs), developmental (time to metamorphosis, weight and length at various developmental stages, protein profile of adult globin) and genetic (adult globin mRNA levels) measures. However, we were unable to detect any differences between treated and control animals. Further, globin delivery using Bohle iridovirus, an Australian ranavirus isolate belonging to the Iridovirus family, did not reduce the survival of metamorphs or alter the form of beta globin expressed in metamorphs.
CONCLUSIONS/SIGNIFICANCE: While we were able to show for the first time that the switch from tadpole to adult globin does occur in B. marinus, we were not able to induce autoimmunity and disrupt metamorphosis. The short development time of B. marinus tadpoles may preclude this approach.
巨蟾蜍, Bufo ( Chaunus ) marinus ,是过去 200 年来引入澳大利亚的最臭名昭著的脊椎动物害虫之一,迄今为止,为了寻找一种可用于生物控制剂的天然存在的巨蟾蜍特异性病原体,人们进行了许多努力,但都没有成功。我们探索了一种替代方法,即对具有广泛宿主特异性的病原体进行基因修饰,使其不再致病,但携带一种基因,以特定于物种的方式破坏巨蟾蜍的生命周期。
方法/主要发现:选择成年β珠蛋白基因作为自身免疫作为控制巨蟾蜍生物控制方法的概念验证的模型基因。先前的一份报告显示,给牛蛙蝌蚪注射成年β珠蛋白会导致变态期表达的β珠蛋白形式发生改变,并降低存活率。在 B. marinus 中,我们首次证实了从蝌蚪到成年珠蛋白的转变。然后,使用行为学(蝌蚪的游泳速度和变态期的跳跃长度)、发育学(变态时间、不同发育阶段的体重和长度、成年珠蛋白的蛋白质谱)和遗传学(成年珠蛋白 mRNA 水平)评估了用纯化的重组成年珠蛋白蛋白注射 B. marinus 蝌蚪的效果。然而,我们未能检测到处理过的和对照组动物之间的任何差异。此外,使用 Bohle 虹彩病毒(属于虹彩病毒科的澳大利亚 Ranavirus 分离株)进行珠蛋白传递并没有降低变态期的存活率,也没有改变变态期表达的β珠蛋白形式。
结论/意义:虽然我们首次证明了 B. marinus 中确实存在从蝌蚪到成年珠蛋白的转变,但我们无法诱导自身免疫并破坏变态。B. marinus 蝌蚪的短暂发育期可能会阻止这种方法的实施。