Interplay between Yersinia pestis and its flea vector in lipoate metabolism

Abstract

To thrive, vector-borne pathogens must survive in the vector’s gut. How these pathogens successfully exploit this environment in time and space has not been extensively characterized. Using Yersinia pestis (the plague bacillus) and its flea vector, we developed a bioluminescence-based approach and employed it to investigate the mechanisms of pathogenesis at an unprecedented level of detail. Remarkably, lipoylation of metabolic enzymes, via the biosynthesis and salvage of lipoate, increases the Y. pestis transmission rate by fleas. Interestingly, the salvage pathway’s lipoate/octanoate ligase LplA enhances the first step in lipoate biosynthesis during foregut colonization but not during midgut colonization. Lastly, Y. pestis primarily uses lipoate provided by digestive proteolysis (presumably as lipoyl peptides) rather than free lipoate in blood, which is quickly depleted by the vector. Thus, spatial and temporal factors dictate the bacterium’s lipoylation strategies during an infection, and replenishment of lipoate by digestive proteolysis in the vector might constitute an Achilles’ heel that is exploited by pathogens.

CsrA Enhances Cyclic-di-GMP Biosynthesis and Yersinia pestis Biofilm Blockage of the Flea Foregut by Alleviating Hfq-Dependent Repression of the hmsT mRNA

“…CsrA mutants in other bacterial species also show slow growth kinetics ( 28 , 58 ). Additionally, the bacterial loads and infection rates of the Δ csrA strain resembled those reported for biofilm-deficient Y. pestis strains ( 7 , 59 – 61 ). Biofilm is thought to maintain bacteria in aggregates that are not easily cleared through defecation after flea blood-feeding and digestion (reviewed in reference 3 ), accounting for lower bacterial number in strains with reduced biofilm levels.…”

Transposon mutagenesis of Rickettsia felis sca1 confers a distinct phenotype during flea infection

“…B) Representative agarose gel of the amplification of adjacent genes downstream of sca1 using PCR from cDNA samples of R. felis sca1::tn (lanes 2, 6, 10) and R. felis WT (lanes 1, 5, 9). Rickettsial DNA samples were used as controls for gene amplification (lanes [13][14][15][16][17][18]. cDNA samples lacking reverse transcriptase were used as a negative control (lanes 3, 4, 7, 8, 11, 12).…”

“…Genetic modification of vector-borne pathogens, such as Yersinia pestis and Bartonella henselae, has identified bacteria-derived factors essential for infection or transmission in fleas [14][15][16][17][18][19][20]. In contrast to these extracellular pathogens, the fastidious nature of rickettsiae requires direct interaction with host cells for propagation, complicating the development of applicable molecular tools.…”

A Widefield Light Microscopy-Based Approach Provides Further Insights into the Colonization of the Flea Proventriculus by Yersinia pestis

“…3D). This difference may be due to the digestion process, which involves the release of enzymes, nutrient-sequestering molecules and bactericidal molecules into the midgut (8,15,16,18). In other words, digestion might generate a bactericidal environment in the midgut.…”

“…none, very few, few, or many) (4). Next, we developed an algorithm that analyzed the fluorescence microscopy images and measured the surface area occupied by fluorescent bacteria within each individual proventriculus (15,16). Although this computerized approach is more accurate than the visual method, it still provides only a rough estimation of the state of proventriculus colonization; only the area occupied by the bacteria is taken into account, and the technique does not consider the fluorescence signal's intensity in the various part of the proventriculus.…”