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Yersinia pestis Δail Mutants Are Not Susceptible to Human Complement Bactericidal Activity in the Flea
Anna M Kolodziejek1, Scott W Bearden2, Sarah Maes2
1Department of Animal, Veterinary and Food Science, University of Idaho, Moscow, Idaho, USA.
Abstract:
Ail confers serum resistance in humans and is a critical virulence factor of Y. pestis, the causative agent of plague. Here, the contribution of Ail for Y. pestis survival in the flea vector was examined. Rat or human but not mouse sera were bactericidal against a Y. pestis Δail mutant at 28°C in vitro. Complement components deposited rapidly on the Y. pestis surface as measured by immunofluorescent microscopy. Ail reduced the amount of active C3b on the Y. pestis surface. Human sera retained bactericidal activity against a Y. pestis Δail mutant in the presence of mouse sera. However, in the flea vector, the serum protective properties of Ail were not required. Flea colonization studies using murine sera and Y. pestis KIM6+ wild type, a Δail mutant, and the Δail/ail control showed no differences in bacterial prevalence or numbers during the early stage of flea colonization. Similarly, flea studies with human blood showed Ail was not required for serum resistance. Finally, a variant of Ail (AilF100V E108_S109insS) from a human serum-sensitive Y. pestis subsp. microtus bv. Caucasica 1146 conferred resistance to human complement when expressed in the Y. pestis KIM6+ Δail mutant. This indicated that Ail activity was somehow blocked, most likely by lipooligosaccharide, in this serum sensitive strain. IMPORTANCE This work contributes to our understanding of how highly virulent Y. pestis evolved from its innocuous enteric predecessor. Among identified virulence factors is the attachment invasion locus protein, Ail, that is required to protect Y. pestis from serum complement in all mammals tested except mice. Murine sera is not bactericidal. In this study, we asked, is bactericidal sera from humans active in Y. pestis colonized fleas? We found it was not. The importance of this observation is that it identifies a protective niche for the growth of serum sensitive and nonsensitive Y. pestis strains.
Insights
The attachment and invasion locus protein (Ail) protects Yersinia pestis from serum, but this protection is not needed for flea colonization. Y. pestis can grow in fleas even when exposed to human serum, suggesting fleas provide a protective niche.
Area of Science:
- Microbiology
- Immunology
- Vector-borne diseases
Background:
- * Yersinia pestis, the plague-causing bacterium, possesses the attachment and invasion locus (Ail) protein, a key virulence factor conferring resistance to mammalian serum.
- * Ail is known to protect Y. pestis from complement-mediated bactericidal activity in humans and other mammals, except mice, whose serum is not bactericidal.
- * The role of Ail in Y. pestis survival within the flea vector, a critical component of plague transmission, remained largely unexplored.
Purpose of the Study:
- * To investigate the contribution of the Ail protein to Y. pestis survival and serum resistance within the flea vector.
- * To determine if the serum-protective properties of Ail are essential for Y. pestis colonization of fleas, particularly when exposed to human or murine sera.
- * To understand the evolutionary implications of Ail's function in the context of Y. pestis transmission dynamics.
Main Methods:
- * Comparative analysis of wild-type Y. pestis and an Δail mutant in serum bactericidal assays *in vitro* using human, rat, and mouse sera.
- * Immunofluorescence microscopy to visualize complement component deposition (C3b) on bacterial surfaces.
- * Flea colonization experiments using Y. pestis strains (wild-type, Δail mutant, and complemented strain) exposed to murine and human blood/sera.
- * Functional analysis of a variant Ail protein from a serum-sensitive strain expressed in a serum-resistant background.
Main Results:
- * *In vitro*, human and rat sera were bactericidal against the Y. pestis Δail mutant, while mouse serum was not, consistent with Ail's role in complement resistance.
- * Ail was found to reduce active C3b deposition on the bacterial surface.
- * Crucially, Ail's serum-protective function was dispensable for Y. pestis colonization in fleas, irrespective of whether murine or human blood was used.
- * A variant Ail protein conferred complement resistance, suggesting that in some strains, Ail's activity might be inhibited, possibly by lipooligosaccharides.
Conclusions:
- * The flea vector provides a protective niche for Y. pestis, rendering the serum resistance conferred by Ail unnecessary for early colonization.
- * This finding highlights that virulence factors crucial for mammalian hosts may not be essential in the arthropod vector.
- * The study suggests that serum-sensitive and serum-nonsensitive Y. pestis strains can potentially thrive in fleas, impacting plague transmission dynamics and evolution.
