Iatsenko Igor, Boichenko Iuliia, Sommer Ralf J
Max Planck Institute for Developmental Biology, Department of Evolutionary Biology, Tübingen, Germany.
Appl Environ Microbiol. 2014 May;80(10):3266-75. doi: 10.1128/AEM.00464-14. Epub 2014 Mar 14.
Bacillus thuringiensis has been widely used as a biopesticide, primarily for the control of insect pests, but some B. thuringiensis strains specifically target nematodes. However, nematicidal virulence factors of B. thuringiensis are poorly investigated. Here, we describe virulence factors of nematicidal B. thuringiensis DB27 using Caenorhabditis elegans as a model. We show that B. thuringiensis DB27 kills a number of free-living and animal-parasitic nematodes via intestinal damage. Its virulence factors are plasmid-encoded Cry protoxins, since plasmid-cured derivatives do not produce Cry proteins and are not toxic to nematodes. Whole-genome sequencing of B. thuringiensis DB27 revealed multiple potential nematicidal factors, including several Cry-like proteins encoded by different plasmids. Two of these proteins appear to be novel and show high similarity to Cry21Ba1. Named Cry21Fa1 and Cry21Ha1, they were expressed in Escherichia coli and fed to C. elegans, resulting in intoxication, intestinal damage, and death of nematodes. Interestingly, the effects of the two protoxins on C. elegans are synergistic (synergism factor, 1.8 to 2.5). Using purified proteins, we determined the 50% lethal concentrations (LC50s) for Cry21Fa1 and Cry21Ha1 to be 13.6 μg/ml and 23.9 μg/ml, respectively, which are comparable to the LC50 of nematicidal Cry5B. Finally, we found that signaling pathways which protect C. elegans against Cry5B toxin are also required for protection against Cry21Fa1. Thus, B. thuringiensis DB27 produces novel nematicidal protoxins Cry21Fa1 and Cry21Ha1 with synergistic action, which highlights the importance of naturally isolated strains as a source of novel toxins.
苏云金芽孢杆菌已被广泛用作生物杀虫剂，主要用于防治害虫，但一些苏云金芽孢杆菌菌株专门针对线虫。然而，苏云金芽孢杆菌的杀线虫毒力因子研究较少。在此，我们以秀丽隐杆线虫为模型描述了杀线虫苏云金芽孢杆菌DB27的毒力因子。我们发现苏云金芽孢杆菌DB27通过肠道损伤杀死多种自由生活和动物寄生线虫。其毒力因子是质粒编码的Cry原毒素，因为质粒消除衍生物不产生Cry蛋白且对线虫无毒。苏云金芽孢杆菌DB27的全基因组测序揭示了多个潜在的杀线虫因子，包括由不同质粒编码的几种类Cry蛋白。其中两种蛋白似乎是新的，与Cry21Ba1具有高度相似性。命名为Cry21Fa1和Cry21Ha1，它们在大肠杆菌中表达并喂食给秀丽隐杆线虫，导致线虫中毒、肠道损伤和死亡。有趣的是，这两种原毒素对秀丽隐杆线虫的作用是协同的（协同因子为1.8至2.5）。使用纯化蛋白，我们确定Cry21Fa1和Cry21Ha1的50%致死浓度（LC50）分别为13.6μg/ml和23.9μg/ml，这与杀线虫Cry5B的LC50相当。最后，我们发现保护秀丽隐杆线虫抵御Cry5B毒素的信号通路对于抵御Cry21Fa1也同样需要。因此，苏云金芽孢杆菌DB27产生具有协同作用的新型杀线虫原毒素Cry21Fa1和Cry21Ha1，这突出了天然分离菌株作为新型毒素来源的重要性。
