Detection of Rickettsia rickettsii, Rickettsia parkeri, and Rickettsia akari in skin biopsy specimens using a multiplex real-time polymerase chain reaction assay.
BACKGROUND
Rickettsia rickettsii, Rickettsia parkeri, and Rickettsia akari are the most common causes of spotted fever group rickettsioses indigenous to the United States. Infected patients characteristically present with a maculopapular rash, often accompanied by an inoculation eschar. Skin biopsy specimens are often obtained from these lesions for diagnostic evaluation. However, a species-specific diagnosis is achieved infrequently from pathologic specimens because immunohistochemical stains do not differentiate among the causative agents of spotted fever group rickettsiae, and existing polymerase chain reaction (PCR) assays generally target large gene segments that may be difficult or impossible to obtain from formalin-fixed tissues.


METHODS
This work describes the development and evaluation of a multiplex real-time PCR assay for the detection of these 3 Rickettsia species from formalin-fixed, paraffin-embedded (FFPE) skin biopsy specimens.


RESULTS
The multiplex PCR assay was specific at discriminating each species from FFPE controls of unrelated bacterial, viral, protozoan, and fungal pathogens that cause skin lesions, as well as other closely related spotted fever group Rickettsia species.


CONCLUSIONS
This multiplex real-time PCR demonstrates greater sensitivity than nested PCR assays in FFPE tissues and provides an effective method to specifically identify cases of Rocky Mountain spotted fever, rickettsialpox, and R. parkeri rickettsiosis by using skin biopsy specimens.
Division of High-Consequence Pathogens and Pathology, Infectious Diseases Pathology Branch.
Amy M
AM
Denison
Department of Pathology, Division of Surgical Pathology, Montefiore Medical Center, Bronx, New York.
Bijal D
BD
Amin
Division of Vector-Borne Diseases, Rickettsial Zoonoses Branch, Centers for Disease Control and Prevention, Atlanta, Georgia.
William L
WL
Nicholson
Division of High-Consequence Pathogens and Pathology, Infectious Diseases Pathology Branch.
Christopher D
CD
Paddock
D001706:Biopsy; D002950:Citrate (si)-Synthase / Q000235:genetics; D005076:Exanthema / Q000382:microbiology*; D006801:Humans; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods; D012281:Rickettsia / Q000235:genetics / Q000302:isolation & purification*; D012282:Rickettsia Infections / Q000175:diagnosis* / Q000382:microbiology / Q000473:pathology; D041141:Rickettsia akari / Q000235:genetics / Q000302:isolation & purification; D012284:Rickettsia rickettsii / Q000235:genetics / Q000302:isolation & purification; D012373:Rocky Mountain Spotted Fever / Q000175:diagnosis* / Q000382:microbiology / Q000473:pathology; D012680:Sensitivity and Specificity; D012867:Skin / Q000382:microbiology / Q000473:pathology
D002950:Citrate (si)-Synthase
Rickettsia; real-time PCR; skin biopsies