Abstract
BackgroundPhlebotomine insects harbor bacterial, viral and parasitic pathogens that can cause diseases of public health importance. Lutzomyia longipalpis is the main vector of visceral leishmaniasis in the New World. Insects can mount a powerful innate immune response to pathogens. Defensin peptides take part in this response and are known to be active against Gram-positive and Gram-negative bacteria, and some parasites. We studied the expression of a defensin gene from Lutzomyia longipalpis to understand its role in sand fly immune response.MethodsWe identified, sequenced and evaluated the expression of a L. longipalpis defensin gene by semi-quantitative RT-PCR. The gene sequence was compared to other vectors defensins and expression was determined along developmental stages and after exposure of adult female L. longipalpis to bacteria and Leishmania.ResultsPhylogenetic analysis showed that the L. longipalpis defensin is closely related to a defensin from the Old World sand fly Phlebotomus duboscqi. Expression was high in late L4 larvae and pupae in comparison to early larval stages and newly emerged flies. Defensin expression was modulated by oral infection with bacteria. The Gram-positive Micrococcus luteus induced early high defensin expression, whilst the Gram-negative entomopathogenic Serratia marcescens induced a later response. Bacterial injection also induced defensin expression in adult insects. Female sand flies infected orally with Leishmania mexicana showed no significant difference in defensin expression compared to blood fed insects apart from a lower defensin expression 5 days post Leishmania infection. When Leishmania was introduced into the hemolymph by injection there was no induction of defensin expression until 72 h later.ConclusionsOur results suggest that L. longipalpis modulates defensin expression upon bacterial and Leishmania infection, with patterns of expression that are distinct among bacterial species and routes of infection.
Highlights
Phlebotomine insects harbor bacterial, viral and parasitic pathogens that can cause diseases of public health importance
Defensin gene sequence and phylogeny The LlDef1 sequence was shown to contain 1034 nucleotides with the coding region between nucleotides 512 and 837 and an intron located between nucleotides 617 and 681 (Figure 1A)
Multiple alignment analysis indicated conserved regions among all defensin sequences selected from blood feeding arthropods and the phylogenetic analysis showed that the L. longipalpis defensin sequence is closely related to defensins obtained from P. duboscqi and other nematocerans (Figure 1B)
Summary
Phlebotomine insects harbor bacterial, viral and parasitic pathogens that can cause diseases of public health importance. Defensin peptides take part in this response and are known to be active against Gram-positive and Gram-negative bacteria, and some parasites. We studied the expression of a defensin gene from Lutzomyia longipalpis to understand its role in sand fly immune response. Lutzomyia longipalpis is the main vector of Leishmania infantum chagasi, the causative agent of visceral leishmaniasis in South America [2]. AMPs have been described in many insects as having a central role in innate immune responses against bacterial and parasitic infections [21,22,23]. We identified, sequenced and investigated the expression profile of a L. longipalpis defensin throughout the sand fly developmental stages, after Gram-positive and Gram-negative bacterial challenges and after sand fly infection with Leishmania mexicana
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