Does transit through the flea vector preadapt Y. pestis to resist mammalian innate immunity? 
When Y. pestis is transmitted into the dermis by an infected flea, it is immediately exposed to the mammalian innate immune system.
The most important antiphagocytic virulence factors, the cytotoxic Yersinia outer proteins (Yops), part of the T3SS encoded by the Y. pestis virulence plasmid and the F1 capsule encoded by the pMT1 plasmid, are not present at this initial stage of infection.
Their expression is strictly temperature-regulated and are not produced in vivo until 3-5 hours after the temperature shift to 37degreesC that accompanies transmission [1],[3],[53],[54].
Consequently, Y. pestis grown at <28degreesC in vitro are initially susceptible to in vivo uptake and killing by phagocytes until the Yop and F1 virulence factors are produced, effectively preventing further phagocytosis [53],[54].
Our results indicate that Y. pestis entering the mammal from an infective flea is relatively resistant to macrophages, as well as PMNs [7]; a vector-specific phenotype that is not related to the T3SS or capsule.
Coming from the flea, Y. pestis is also associated with the biofilm ECM, identical or closely related to the poly-beta-1,6-N-acetyl glucosamine ECM of staphylococcal biofilms, which has been shown to provide protection from innate immune components [55],[56].
In addition, although the antiphagocytic F1 capsule and Psa fimbriae do not appear to be produced in the flea, upregulation in the flea of most F1 genes in the cafRcaf1M1A1 locus and the Psa usher protein gene psaC (Tables 1, S1) suggests that components of the F1 and Psa translocation system are made, which may prime Y. pestis for rapid secretion of these extracellular virulence factors after transmission.
The upregulation of the innate immunity resistance genes phoP and mgtC suggest that those Y. pestis that are phagocytized may be prepared for resistance to CAMPs and intracellular survival while still in the flea vector.
Finally, the major essential virulence factors yadBC and pla, essential for Y. pestis dissemination from the dermis, were maximally or very highly expressed in the flea (Tables 1, S3).
Besides degrading plasminogen, the Pla protease may also inactivate CAMPs, particularly when the F1 capsule is not present [57], which matches the phenotype of Y. pestis in the flea.
In summary, Y. pestis appears to be prepared for pathogenesis in the mammal while still in the flea vector.
The biofilm phenotype of Y. pestis and the virulence factors upregulated or highly expressed in the flea may enhance the earliest stages of plague pathogenesis while the full complement of temperature-shift-regulated virulence factors is still being induced.
Increased resistance to innate immunity that is preinduced in the flea vector may be critical to productive transmission because blocked fleas transmit relatively few bacteria, often below the LD50 of Y. pestis grown in vitro at <28degreesC [1],[52].
