Dirofilaria immitis is the globally distributed agent of heartworm disease. Infection in canines causes debilitating disease that can be fatal if left untreated. Macrocyclic lactones can prevent heartworm disease in dogs, cats and ferrets by killing larvae before they develop into adult worms in the pulmonary artery. However, administration of prophylactic drugs to wild canids to prevent D. immitis infection is not feasible. Furthermore, a vaccine against heartworm is currently unavailable and drug resistant D. immitis have been identified, highlighting the need for new strategies to prevent parasite transmission. We recently established a method to block development of emerging third-stage larvae (eL3) from the mosquito Aedes aegypti by over-activating the Toll pathway, one of the major innate immune signaling pathways in mosquitoes. Our previous study used a drug-sensitive strain of D. immitis and it remains unknown if the strategy is effective against different D. immitis genotypes and, more importantly, if it would work against drug-resistant genotypes. The purpose of this study was to determine whether Toll pathway activation is capable of blocking eL3 development of D. immitis strains that are resistant to macrocyclic lactones. We infected mosquitoes with two independent strains of D. immitis previously confirmed as being resistant to macrocyclic lactones, and then activated Toll signaling by RNAi-mediated silencing of the pathway inhibitor, IκB/Cactus, and quantitatively measured eL3 development. Similar to the drug-sensitive strain, eL3 were strongly reduced by Toll activation in both drug-resistant strains. Furthermore, similar to the drug-sensitive strain, the reduction of eL3 in both drug-resistant strains suggests a defect in larval invasion of, or development in, the Malpighian tubules – the organ in the mosquito to which microfilariae migrate after ingestion and where the larvae undergo several developmental molts. In summary, Toll pathway activation blocks the development of three distinct D. immitis genotypes, including two different drug-resistant genotypes. If this strategy can be applied to heartworm vectors in the field, it may help reduce the spread of disease and is not predicted to favor the spread of drug resistance.

Dirofilaria炎是丝虫病的全球传播媒介。犬类感染会导致衰弱性疾病，如果不及时治疗，可能会致命。大环内酯可通过杀死幼虫，使其在肺动脉中成虫之前杀死幼虫，从而预防狗，猫和雪貂的心丝虫病。但是，向野犬施用预防药物以预防D. Imm炎感染是不可行的。此外，目前尚没有针对丝虫的疫苗，并且已经鉴定出了耐药性的D. immitis，这突出表明需要采取新策略来预防寄生虫传播。我们最近建立了一种方法来阻止蚊子埃及伊蚊的第三阶段幼虫（eL3）的发育通过过度激活Toll途径，Toll途径是蚊子的主要先天免疫信号传导途径之一。我们先前的研究使用的是对D. immitis药物敏感的菌株，目前尚不清楚该策略是否对不同的D. immitis基因型有效，更重要的是，该策略是否对耐药性基因型有效。这项研究的目的是确定Toll途径激活是否能够阻断对大环内酯有抗性的D.炎性菌株的eL3发育。我们用两种独立的D. Immitis菌株感染了蚊子先前证实对大环内酯具有抗性，然后通过RNAi介导的途径抑制剂IκB/仙人掌沉默和定量测量的eL3发育激活了Toll信号转导。与药物敏感菌株相似，两种耐药菌株中的Toll激活均会强烈降低eL3。此外，与药物敏感菌株相似，两种耐药菌株中eL3的减少均表明幼虫侵袭或发展了Malpighian小管，该小管是蚊子中的器官，吸食后微丝aria会迁移到该器官，幼虫经历了几次蜕皮。总之，Toll通路的激活阻碍了三种截然不同的D. Immitis的发展基因型，包括两种不同的耐药基因型。如果该策略可以在野外应用于丝虫载体，则可能有助于减少疾病的传播，并且预计不会有利于耐药性的传播。