Model of early-phase regurgitative transmission of <i>Y</i>. <i>pestis</i> by PIER positive and PIER negative fleas.

<p>Following an infectious blood meal, fleas rapidly lyse red blood cells (RBC) and hemoglobin is released. Within a few hours, <i>Y</i>. <i>pestis</i> coalesces into aggregates that are enveloped in an exogenous matrix that is derived from blood digestion products [<a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1006859#ppat.1006859.ref008" target="_blank">8</a>]. Depending on the blood source, incompletely digested RBCs and oxyhemoglobin crystals can associate with the bacterial aggregates in the midgut. Normal peristalsis and proventricular pulsations, which continue long after feeding, draw the aggregates into the proventriculus, and some of them lodge there. According to the model, when fleas with extensive proventricular colonization feed again 1–3 days after infection, the incoming blood flow is sufficiently impeded to result in regurgitation of bacteria. Early proventricular colonization and early-phase transmission do not require mature biofilm formation and the permanent proventricular colonization that is mediated by the <i>Y</i>. <i>pestis hmsHFRS</i> genes [<a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1006859#ppat.1006859.ref008" target="_blank">8</a>, <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1006859#ppat.1006859.ref015" target="_blank">15</a>]. (A) In PIER-negative fleas, RBCs are digested quickly and hemoglobin remains soluble. The bacterial aggregates in the proventriculus of heavily colonized fleas withstand the initial pulses of incoming blood, which can result in regurgitative transmission of a few bacteria. However, the obstruction is ephemeral and incoming blood quickly dislodges most or all of the poorly adherent bacterial aggregates back into the midgut. The proventriculus can later be recolonized permanently by an Hms-dependent biofilm. (B) In PIER-positive fleas, less rapid RBC digestion and formation of oxyhemoglobin crystals correlate with a more tenacious, expansive proventricular colonization and extrusion of infected blood into the esophagus (post-infection esophageal reflux; PIER). The obstruction in the PV is more resistant to dislodgement and bacteria are already present in the esophagus, both of which enhance regurgitative EPT. Incoming blood ultimately channels through the proventricular obstruction of most fleas and enters the midgut. However, because PIER tends to stabilize the initial proventricular colonization, the extrinsic incubation period for biofilm-dependent transmission (BDT) may be shortened, and the two phases of transmission may temporally overlap in populations of infected fleas. Figure adapted from reference [<a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1006859#ppat.1006859.ref008" target="_blank">8</a>].