The Cat Flea (Ctenocephalides felis) Immune Deficiency Signaling Pathway Regulates Rickettsia typhi Infection

Abstract

Rickettsia species are obligate intracellular bacteria with both conserved and lineage-specific strategies for invading and surviving within eukaryotic cells. One variable component of Rickettsia biology involves arthropod vectors: for instance, typhus group rickettsiae are principally vectored by insects (i.e., lice and fleas), whereas spotted fever group rickettsiae are exclusively vectored by ticks. For flea-borne Rickettsia typhi, the etiological agent of murine typhus, research on vertebrate host biology is facilitated using cell lines and animal models. However, due to the lack of any stable flea cell line or a published flea genome sequence, little is known regarding R. typhi biology in flea vectors that, importantly, do not suffer lethality due to R. typhi infection. To address if fleas combat rickettsial infection, we characterized the cat flea (Ctenocephalides felis) innate immune response to R. typhi. Initially, we determined that R. typhi infects Drosophila cells and increases antimicrobial peptide (AMP) gene expression, indicating immune pathway activation. While bioinformatics analysis of the C. felis transcriptome identified homologs to all of the Drosophila immune deficiency (IMD) and Toll pathway components, an AMP gene expression profile in Drosophila cells indicated IMD pathway activation upon rickettsial infection. Accordingly, we assessed R. typhi-mediated flea IMD pathway activation in vivo using small interfering RNA (siRNA)-mediated knockdown. Knockdown of Relish and Imd increased R. typhi infection levels, implicating the IMD pathway as a critical regulator of R. typhi burden in C. felis. These data suggest that targeting the IMD pathway could minimize the spread of R. typhi, and potentially other human pathogens, vectored by fleas.

Rickettsia Lipid A Biosynthesis Utilizes the Late Acyltransferase LpxJ for Secondary Fatty Acid Addition

“…Primers used in this study were obtained from Integrated DNA Technologies (IDT) and are listed in Table 2 . Genomic DNA was isolated from R. typhi grown in Vero76 tissue monolayer as previously described ( 53 ). lpxJ Rt (RT0047; NCBI accession number AAU03534 ) was amplified using Q5 polymerase 2× master mix (M0492; NEB) and gel purified (740609.50; Macherey-Nagel) before infusion cloning (639649; Clontech) into a lac -inducible pFLAG-ctc vector (discontinued; Sigma) using HindIII and XhoI restriction sites.…”

Transcriptomic profiling of the digestive tract of the rat flea, Xenopsylla cheopis, following blood feeding and infection with Yersinia pestis

“…Drosophila and Galleria mellonella have recently been adapted as models for Y. pestisinsect interactions and have been used to show that the bacterial PhoPQ two-component gene regulatory system and the OxyR transcription factor mediate resistance to AMPs and ROS and are required for colonization of Drosophila and Galleria larvae, as well as the rat flea gut [8,17,18]. In studies of flea physiology and genetics, primarily with the cat flea, Ctenocephalides felis, many canonical insect responses to pathogens have been observed [7,[19][20][21][22]. Recent studies using cultured Drosophila cells and RNA interference in cat fleas indicate that R. typhi infection is moderated by antimicrobials generated by the immune deficiency (IMD) pathway, the major insect innate immune cascade that detects and responds to DAP-type peptidoglycan, common in Gram-negative bacteria [19].…”

“…In studies of flea physiology and genetics, primarily with the cat flea, Ctenocephalides felis, many canonical insect responses to pathogens have been observed [7,[19][20][21][22]. Recent studies using cultured Drosophila cells and RNA interference in cat fleas indicate that R. typhi infection is moderated by antimicrobials generated by the immune deficiency (IMD) pathway, the major insect innate immune cascade that detects and responds to DAP-type peptidoglycan, common in Gram-negative bacteria [19]. Furthermore, transcriptomic data indicate that the IMD pathway is conserved between C. felis and the Diptera [19].…”

“…pestis -insect interactions and have been used to show that the bacterial PhoPQ two-component gene regulatory system and the OxyR transcription factor mediate resistance to AMPs and ROS and are required for colonization of Drosophila and Galleria larvae, as well as the rat flea gut [ 8 , 17 , 18 ]. In studies of flea physiology and genetics, primarily with the cat flea, Ctenocephalides felis , many canonical insect responses to pathogens have been observed [ 7 , 19 – 22 ]. Recent studies using cultured Drosophila cells and RNA interference in cat fleas indicate that R .…”

Annexin A2 depletion exacerbates the intracerebral microhemorrhage induced by acute rickettsia and Ebola virus infections

“…Rickettsioses represent devastating human infections [1][2][3][4][5][6][7]. These arthropod-borne diseases are caused by obligatory intracellular bacteria of the genus Rickettsia (R.).…”