Rickettsia parkeri with a Genetically Disrupted Phage Integrase Gene Exhibits Attenuated Virulence and Induces Protective Immunity against Fatal Rickettsioses in Mice

Esteban Arroyave,Rong Fang,Yong-Fang Kuo,Tian Wang,Ulrike Munderloh,Nicole Burkhardt,Ilirjana Hyseni

doi:10.3390/pathogens10070819

Abstract

Although rickettsiae can cause life-threatening infections in humans worldwide, no licensed vaccine is currently available. To evaluate the suitability of live-attenuated vaccine candidates against rickettsioses, we generated a Rickettsia parkeri mutant RPATATE_0245::pLoxHimar (named 3A2) by insertion of a modified pLoxHimar transposon into the gene encoding a phage integrase protein. For visualization and selection, R. parkeri 3A2 expressed mCherry fluorescence and resistance to spectinomycin. Compared to the parent wild type (WT) R. parkeri, the virulence of R. parkeri 3A2 was significantly attenuated as demonstrated by significantly smaller size of plaque, failure to grow in human macrophage-like cells, rapid elimination of Rickettsia and ameliorated histopathological changes in tissues in intravenously infected mice. A single dose intradermal (i.d.) immunization of R. parkeri 3A2 conferred complete protection against both fatal R. parkeri and R. conorii rickettsioses in mice, in association with a robust and durable rickettsiae-specific IgG antibody response. In summary, the disruption of RPATATE_0245 in R. parkeri resulted in a mutant with a significantly attenuated phenotype, potent immunogenicity and protective efficacy against two spotted fever group rickettsioses. Overall, this proof-of-concept study highlights the potential of R. parkeri mutants as a live-attenuated and multivalent vaccine platform in response to emergence of life-threatening spotted fever rickettsioses.

Highlights

In recent years, an increasing number of cases of rickettsial disease has been reported in several regions of North America where rickettsioses are endemic as well as in areas where rickettsioses have not commonly been diagnosed [1,2]
Because the transposon included the aadA gene for selection, we tested whether R. parkeri Rickettsia parkeri mutant RPATATE_0245 (3A2) expressed antibiotic resistance (Figure 2A–C)
In the regular growth medium, R. parkeri 3A2 replicated at a rate similar to R. parkeri wild type (WT) over three days (Figure 2A) (p = 0.5366 by Friedman test)

Summary

Introduction

An increasing number of cases of rickettsial disease has been reported in several regions of North America where rickettsioses are endemic as well as in areas where rickettsioses have not commonly been diagnosed [1,2]. Life-threatening forms of spotted fevers (case fatality rate of 5–10% or more) are caused by Rickettsia rickettsii that is transmitted by wood ticks (Dermacentor andersoni and Dermacentor variabilis) in the Northern Rocky Mountains and South Central States, respectively, and brown dog ticks (Rhipicephalus sanguineus) in Arizona and neighboring regions of the USA [3]. In South America, the incidence of Brazilian spotted fever has increased significantly, disproportionately affecting rural populations with poor access to health care services [4]. The Brazilian strain of R. rickettsii is transmitted by Amblyomma ticks [5], and the mortality rate is very high at 50% [6]. Due to the high case fatality rate, susceptibility of humans to infection via the aerosol route, and potential use as a bioweapon [7], an effective vaccine is urgently needed, but an FDA approved, and licensed vaccine is not available

Methods

Results

Conclusion