The Role of Antimicrobial Peptides in the Innate Immunity of Insects and Their Mode of Action
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Kaushal, Akanksha
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Abstract
Antimicrobial peptides (AMPs) are components of both vertebrate and invertebrate innate immune systems that are expressed in response to exposure to bacterial antigens. Naturally occurring AMPs from evolutionarily ancient species have been extensively studied and are being developed as potential therapeutics against antibiotic resistant microorganisms. In this thesis, a putative Cimex lectularius (bedbug, CL) defensin is characterized for its effectiveness against human skin flora including Gram-negative and Gram-positive bacteria. The bedbug defensin (CL-defensin), belonging to family of insect defensins, is predicted to have a characteristic N-terminal loop, an α-helix, and an antiparallel β-sheet, which was supported by circular dichroism spectroscopy. The defensin was shown to be antimicrobial against Gram-positive bacteria commonly found on human skin (Micrococcus luteus, Corynebacterium renale, Staphylococcus aureus and Staphylococcus epidermidis); however, it was ineffective against common skin xii Gram-negative bacteria (Pseudomonas aeruginosa and Acinetobactor baumannii) under low-salt conditions. CL-defensin was also effective against Micrococcus luteus and Corynebacterium renale in high-salt (MIC) conditions. Our studies indicate that CL-defensin functions by depolarization and pore-formation in the bacterial cytoplasmic membrane. In addition to bedbugs, this thesis also focuses on characterizing antimicrobial activity of mosquito Aedes albopitctus’ antimicrobial peptides against Francisella. Francisella tularensis is the cause of the zoonotic disease tularemia. In Sweden and Scandinavia, epidemiological studies have implicated mosquitoes as a vector. Prior research has demonstrated the presence of Francisella DNA in infected mosquitoes but has not shown transmission of tularemia from a mosquito to a mammalian host. We hypothesized that antimicrobial peptides may play a role in mosquito host-defense to Francisella. We established that Francisella sp. are susceptible to two Cecropin antimicrobial peptides derived from the mosquito Aedes albopictus. We also demonstrated induced gene expression of these peptides by Francisella infection C6/36 mosquito cell line. We demonstrated that mosquito antimicrobial peptides are active against Francisella by disrupting the cellular membrane of the bacteria. Thus, antimicrobial peptides may play a role in the inability of mosquitoes to establish an effective natural transmission of tularemia.
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This thesis has been embargoed for 5 years. It will not be available until April 2021 at the earliest.
Keywords
Defensin, Antimicrobial peptide, EC50, Pore formation, Depolarization