ABSTRACT

Mammalian infection models have contributed significantly to our understanding of the host-mycobacterial interaction, revealing potential mechanisms and targets for novel antimycobacterial therapeutics. However, the use of conventional mammalian models such as mice, are typically expensive, high maintenance, require specialized animal housing, and are ethically regulated. Furthermore, research using Mycobacterium tuberculosis (MTB), is inherently difficult as work needs to be carried out at biosafety level 3 (BSL3). The insect larvae of Galleria mellonella (greater wax moth), have become increasingly popular as an infection model, and we previously demonstrated its potential as a mycobacterial infection model using Mycobacterium bovis BCG. Here we present a novel BSL2 complaint MTB infection model using G. mellonella in combination with a bioluminescent ΔleuDΔpanCD double auxotrophic mutant of MTB H37Rv (SAMTB lux) which offers safety and practical advantages over working with wild type MTB. Our results show a SAMTB lux dose dependent survival of G. mellonella larvae and demonstrate proliferation and persistence of SAMTB lux bioluminescence over a 1 week infection time course. Histopathological analysis of G. mellonella, highlight the formation of early granuloma-like structures which matured over time. We additionally demonstrate the drug efficacy of first (isoniazid, rifampicin, and ethambutol) and second line (moxifloxacin) antimycobacterial drugs. Our findings demonstrate the broad potential of this insect model to study MTB infection under BSL2 conditions. We anticipate that the successful adaptation and implementation of this model will remove the inherent limitations of MTB research at BSL3 and increase tuberculosis research output.

摘要

哺乳动物感染模型为我们对宿主-分枝杆菌相互作用的理解做出了重要贡献，揭示了新型抗分枝杆菌疗法的潜在机制和靶标。但是，使用常规的哺乳动物模型，例如小鼠，通常价格昂贵，维护成本高，需要专门的动物住房并且受到道德规范。此外，使用结核分枝杆菌（MTB）进行研究固有地困难，因为需要在生物安全等级3（BSL3）下进行工作。圆盘虫（大蜡蛾）的昆虫幼虫已越来越受感染模型的欢迎，并且我们之前已经证明了其使用牛分枝杆菌作为分枝杆菌感染模型的潜力卡介苗。在这里，我们介绍了一种新的BSL2投诉MTB感染模型，它使用了G. mellonella与MTB H37Rv（SAMTB lux）的生物发光ΔleuDΔpanCD双营养缺陷型突变体相结合，与使用野生型MTB相比，它具有安全性和实用性。我们的研究结果显示SAMTB勒克斯的剂量依赖性生存螟幼虫和展示增殖和SAMTB的持续勒克斯生物发光超过1周的感染时间过程。mel.ella mellonella的组织病理学分析，突出了早期肉芽肿样结构的形成，并随着时间的推移逐渐成熟。我们还证明了第一类（异烟肼，利福平和乙胺丁醇）和第二类（莫西沙星）抗分枝杆菌药物的药效。我们的发现证明了这种昆虫模型在BSL2条件下研究MTB感染的广泛潜力。我们预计该模型的成功适应和实施将消除BSL3上MTB研究的固有局限性，并增加结核病研究成果。