Amblyomma imitator Ticks as Vectors of Rickettsia rickettsii, Mexico

Real-time PCR of Amblyomma imitator tick egg masses obtained in Nuevo Leon State, Mexico, identified a Rickettsia species. Sequence analyses of 17-kD common antigen and outer membrane protein A and B gene fragments showed to it to be R. rickettsii, which suggested a potential new vector for this bacterium

Experimental Infection of Opossums Didelphis aurita by Rickettsia rickettsii and Evaluation of the Transmission of the Infection to Ticks Amblyomma cajennense

The present study evaluated the infection of opossums (Didelphis aurita) by Rickettsia rickettsii and their role as amplifier hosts for horizontal transmission of R. rickettsii to Amblyomma cajennense ticks. Three groups of opossums were evaluated: on day 0, group 1 (G1) was inoculated intraperitoneally with R. rickettsii; group 2 (G2) was infested by R. rickettsii-infected ticks; and group 3 (G3) was the uninfected control group. Opossum rectal temperature was measured daily. Blood samples were collected every 2 to 4 days during 30 days, and used to (1) inoculate guinea pigs intraperitoneally; (2) extract DNA followed by real-time polymerase chain reaction (PCR) targeting the rickettsial gene gltA; (3) study hematology; (4) detect R. rickettsii-reactive antibodies by indirect direct immunofluorescence assay (IFA). Blood was also collected every 10 days from days 30 to 180, to be tested by serology. Opossums were infested by uninfected A. cajennense larvae and nymphs from days 3 to 15. Engorged ticks were collected and allowed to molt in an incubator. Thereafter, the subsequent flat ticks were allowed to feed on uninfected rabbits, which were tested for seroconversion by IFA. Samples of flat ticks were also tested by real-time PCR. All G1 and G2 opossums became infected by R. rickettsii, as demonstrated by real-time PCR or/and guinea pig inoculation, but they showed no clinical abnormality. Rickettsemia was first detected at days 2 to 8, lasting intermittently till days 1 to 30. Approximately 18% and 5% of the flat ticks previously fed on G1 and G2 opossums, respectively, became infected by R. rickettsii, but only the rabbits infested with G1-derived ticks seroconverted. The study demonstrated that R. rickettsii was capable of infecting opossums without causing illness and developing rickettsemia capable of causing infection in guinea pigs and ticks, although the infection rate in ticks was low

Hematogenously disseminated Orientia tsutsugamushi-infected murine model of scrub typhus [corrected].

Orientia tsutsugamushi, the etiologic agent of scrub typhus, is a mite-borne rickettsia transmitted by the parasitic larval stage of trombiculid mites. Approximately one-third of the world's population is at risk of infection with Orientia tsutsugamushi, emphasizing its importance in global health. In order to study scrub typhus, Orientia tsutsugamushi Karp strain has been used extensively in mouse studies with various inoculation strategies and little success in inducing disease progression similar to that of human scrub typhus. The objective of this project was to develop a disease model with pathology and target cells similar to those of severe human scrub typhus. This study reports an intravenous infection model of scrub typhus in C57BL/6 mice. This mouse strain was susceptible to intravenous challenge, and lethal infection occurred after intravenous inoculation of 1.25 × 10(6) focus (FFU) forming units. Signs of illness in lethally infected mice appeared on day 6 with death occurring ∼ 6 days later. Immunohistochemical staining for Orientia antigens demonstrated extensive endothelial infection, most notably in the lungs and brain. Histopathological analysis revealed cerebral perivascular, lymphohistiocytic infiltrates, focal hemorrhages, meningoencephalitis, and interstitial pneumonia. Disseminated infection of endothelial cells with Orientia in C57BL/6 mice resulted in pathology resembling that of human scrub typhus. The use of this model will allow detailed characterization of the mechanisms of immunity to and pathogenesis of O. tsutsugamushi infection

Canine infection by Rickettsiae and Ehrlichiae in southern Brazil

This study evaluated the infection caused by Rickettsia and Ehrlichia agents among dogs in southern Brazil. A total of 389 dogs were tested by the indirect immunofluorescence assay (IFA) for Rickettsia rickettsii, Rickettsia parkeri, Rickettsia amblyommii, Rickettsia rhipicephali, Rickettsia bellii, and Ehrlichia canis. Overall, 42.4% (165/389) of the dogs were seroreactive to at least one Rickettsia species, but only 11 canine sera reacted with another Rickettsia species without reacting with R. parkeri. A total of 100 (25.7%) canine sera showed titers to R. parkeri at least 4-fold higher than those to any of the other rickettsial antigens, allowing us to consider that these dogs were infected by R. parkeri. Dogs that had direct contact with pasture or forest areas were > 2 times more likely to be seroreactive to Rickettsia than dogs with no such direct contact. Only 19 (4.8%) of the 389 dogs were seroreactive to E. canis

Host EPAC1 Modulates Rickettsial Adhesion to Vascular Endothelial Cells via Regulation of ANXA2 Y23 Phosphorylation

Introduction: Intracellular cAMP receptor exchange proteins directly activated by cAMP 1 (EPAC1) regulate obligate intracellular parasitic bacterium rickettsial adherence to and invasion into vascular endothelial cells (ECs). However, underlying precise mechanism(s) remain unclear. The aim of the study is to dissect the functional role of the EPAC1-ANXA2 signaling pathway during initial adhesion of rickettsiae to EC surfaces. Methods: In the present study, an established system that is anatomically based and quantifies bacterial adhesion to ECs in vivo was combined with novel fluidic force microscopy (FluidFM) to dissect the functional role of the EPAC1-ANXA2 signaling pathway in rickettsiae–EC adhesion. Results: The deletion of the EPAC1 gene impedes rickettsial binding to endothelium in vivo. Rickettsial OmpB shows a host EPAC1-dependent binding strength on the surface of a living brain microvascular EC (BMEC). Furthermore, ectopic expression of phosphodefective and phosphomimic mutants replacing tyrosine (Y) 23 of ANXA2 in ANXA2-knock out BMECs results in different binding force to reOmpB in response to the activation of EPAC1. Conclusions: EPAC1 modulates rickettsial adhesion, in association with Y23 phosphorylation of the binding receptor ANXA2. Underlying mechanism(s) should be further explored to delineate the accurate role of cAMP-EPAC system during rickettsial infection

Bacterial load (47 kDa gene copies/pg of DNA or µL of blood) kinetics of 1.25×10⁴ FFU challenged C57BL/6 mice.

a<p>, ND- not detected.</p><p>*, only two animals remained.</p

Histopathology of the lung following 1.25×10⁴<i>Orientia</i> challenge.

<p>H & E stained uninfected lung tissue (<b>A-20×</b>) and IHC of uninfected lung (<b>B-40×</b>) compared with lung from i.p. inoculated mice 12 dpi (H&E <b>C-20×</b>, IHC <b>D-40×</b>), and lung from i.v. inoculated 15 dpi (H&E <b>E-20×</b>, IHC <b>F-40×</b>). All i.p. infections were lethal with less severe pulmonary cellular infiltrate when compared to that of i.v. infected mice with capillary endothelial cell infection of the aveolar septa.</p

Histopathology of the liver and kidney following lethal intravenous <i>Orientia</i> challenge at days 9 and 12 pi.

<p>Portal triaditis (<b>A-arrow; 20×</b>) was prominent at 9 dpi, and at 12 dpi perivascular infiltrates were observed in the hepatic sinusoids (<b>B-arrow; 20×</b>). Mild perivascular infiltrates were observed in the kidney at 9 dpi (<b>C-20×</b>), and at 12 dpi (<b>D-arrow; 20×</b>) cellular infiltrates were observed throughout the kidney, particularly as peritubular infiltrates.</p

Histopathology of mice following lethal intravenous <i>Orientia</i> challenge at days 9 and 12 pi.

<p>Immunohistochemical staining of tissues from animals 9 and 12(<b>A-20×</b>) was observed in the majority of animals on 9 dpi with cerebral perivascular, lymphohistiocytic infiltrates (<b>B-inset; 40×, C-100×</b>), cerebral hemorrhage (<b>D-100×</b>), and endothelial infection (<b>E-100×</b>) in moribund animals at 11 dpi. Pulmonary cellular infiltrates were marked 9 dpi resulting in interstitial pneumonia (<b>F-20×</b>). At 12 dpi (<b>G-20×</b>), peribronchial and perivascular cellular infiltration, interstitial pneumonia, and edema (<b>G-arrow</b>) were observed in all animals.</p

Bacterial load (47 kDa gene copies/pg of DNA or µL of blood) kinetics of 1.25×10⁶ FFU challenged C57BL/6 mice.

a<p>, ND- not detected.</p