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

Abstract

Yersinia pestis, the causative agent of plague, is a highly lethal pathogen transmitted by the bite of infected fleas. Once ingested by a flea, Y. pestis establish a replicative niche in the gut and produce a biofilm that promotes foregut colonization and transmission. The rat flea Xenopsylla cheopis is an important vector to several zoonotic bacterial pathogens including Y. pestis. Some fleas naturally clear themselves of infection; however, the physiological and immunological mechanisms by which this occurs are largely uncharacterized. To address this, RNA was extracted, sequenced, and distinct transcript profiles were assembled de novo from X. cheopis digestive tracts isolated from fleas that were either: 1) not fed for 5 days; 2) fed sterile blood; or 3) fed blood containing~5x10 8 CFU/ml Y. pestis KIM6+. Analysis and comparison of the transcript profiles resulted in identification of 23 annotated (and 11 unknown or uncharacterized) digestive tract transcripts that comprise the early transcriptional response of the rat flea gut to infection with Y. pestis. The data indicate that production of antimicrobial peptides regulated by the immune-deficiency pathway (IMD) is the primary flea immune response to infection with Y. pestis. The remaining infection-responsive transcripts, not obviously associated with the immune response, were involved in at least one of 3 physiological themes: 1) alterations to chemosensation and gut peristalsis; 2) modification of digestion and metabolism; and 3) production of chitin-binding proteins (peritrophins). Despite producing several peritrophin transcripts shortly after feeding, including a subset that were infection-responsive, no thick peritrophic membrane was detectable by histochemistry or electron microscopy of rat flea guts for the first 24 hours following blood-feeding. Here we discuss the physiological implications of rat flea infection-responsive transcripts, the function of X. cheopis peritrophins, and the mechanisms by which Y. pestis may be cleared from the flea gut.

Molecular and Genetic Mechanisms That Mediate Transmission of Yersinia pestis by Fleas

“…Recently, genomics-based research is beginning to provide more specific insights into the insect host environment of Y. pestis . Transcriptomic profiling analyses revealed a large set of genes that were upregulated in the X. cheopis digestive tract tissue in response to a sterile or Y. pestis -infected blood meal [ 10 ]. Four hours after feeding, upregulated digestive enzyme genes most notably included several that encode serine proteases (trypsins or trypsin-like enzymes) and several enzymes of lipid digestion and metabolism pathways.…”

“…Early after infection with Y. pestis , the expression of several antibacterial effector genes is upregulated in X. cheopis digestive tract epithelia, including those encoding antimicrobial peptides and lysozyme. The response to Y. pestis appears to be due to upregulation of the immune deficiency (Imd) pathway, the arm of insect immunity directed against Gram-negative bacteria [ 10 ]. Another major arm of insect immunity is the dual oxidase-reactive oxygen species (Duox-ROS) system, which generates antibacterial oxygen radicals.…”

“…Prominent virulence factors such as the F1 capsule and pH 6 antigens, iron acquisition operons, and RovA are downregulated [ 25 ]. Transcriptomic analyses of Y. pestis in mammalian host tissue and the flea digestive tract, as well as analyses of the flea transcriptomic response to oral infection with Y. pestis , have pointed out genes and gene regulatory systems that are induced in mammal or flea [ 10 , 13 , 25 , 26 , 27 , 28 , 29 ]. These have helped to lay the groundwork for much current research to identify and characterize the specific molecular interactions of Y. pestis with its insect host that lead to a transmissible infection.…”

“…The flea immune response to oral infection is analogous to the mammalian innate immune response, and the expression of several Y. pestis factors that protect against such a response is upregulated in the flea [ 10 , 27 ]. The PhoPQ two-component system (2CS) senses the decrease in pH experienced in the flea gut, which mediates lipid A modifications that protect against antimicrobial peptides [ 45 ].…”

Yersinia pestislipopolysaccharide remodeling confers resistance to aXenopsylla cheopiscecropin

“…Xenopsylla cheopis, the oriental rat flea, is a major vector for Y. pestis in nature [14][15][16]. Analysis of X. cheopis transcriptomes indicated that different classes of AMPs are expressed in fleas and are responsive to Y. pestis infections [17][18][19]. However, little is known about the antimicrobial activities or mechanisms of action of these peptides.…”