Abstract
Bacterial proteins destined for the Tat pathway are folded before crossing the inner membrane and are typically identified by an N-terminal signal peptide containing a twin arginine motif. Translocation by the Tat pathway is dependent on the products of genes which encode proteins possessing the binding site of the signal peptide and mediating the actual translocation event. In the fully virulent CO92 strain of Yersinia pestis, the tatA gene was deleted. The mutant was assayed for loss of virulence through various in vitro and in vivo assays. Deletion of the tatA gene resulted in several consequences for the mutant as compared to wild-type. Cell morphology of the mutant bacteria was altered and demonstrated a more elongated form. In addition, while cultures of the mutant strain were able to produce a biofilm, we observed a loss of adhesion of the mutant biofilm structure compared to the biofilm produced by the wild-type strain. Immuno-electron microscopy revealed a partial disruption of the F1 antigen on the surface of the mutant. The virulence of the ΔtatA mutant was assessed in various murine models of plague. The mutant was severely attenuated in the bubonic model with full virulence restored by complementation with the native gene. After small-particle aerosol challenge in a pneumonic model of infection, the mutant was also shown to be attenuated. In contrast, when mice were challenged intranasally with the mutant, very little difference in the LD50 was observed between wild-type and mutant strains. However, an increased time-to-death and delay in bacterial dissemination was observed in mice infected with the ΔtatA mutant as compared to the parent strain. Collectively, these findings demonstrate an essential role for the Tat pathway in the virulence of Y. pestis in bubonic and small-aerosol pneumonic infection but less important role for intranasal challenge.
Highlights
Yersinia pestis is the causative agent of plague and primarily a disease of rodents, transmitted typically by fleas
For those clones which reverted back to the wild-type strain, a PCR fragment of 2.1 kb was observed. For those mutants deleted of the tatA gene, a shift of approximately 250 bp was observed
As the deletion of the tatA gene would affect translocated proteins, we examined proteins extracted from both the cell pellet and supernatant fractions of mutant and wild-type bacteria grown in heart infusion (HI) broth at 28uC or 37uC containing CaCl2
Summary
Yersinia pestis is the causative agent of plague and primarily a disease of rodents, transmitted typically by fleas. Clinical forms of the disease in humans are bubonic, septicemic, and pneumonic. Humans most often become infected by fleabites which manifests in the bubonic form of plague and is characterized by painful local lymphadenopathy, referred to as a bubo. Infection of the lymph node may disseminate throughout the body leading to septicemic plague. Y. pestis may reach the lungs and lead to the pneumonic form of plague which could be spread human-tohuman by the respiratory route. Pneumonic plague is the most severe and frequently fatal form of the disease [1,2]
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