Use of real-time quantitative PCR targeting the msp2 protein gene to identify cryptic Anaplasma phagocytophilum infections in wildlife and domestic animals

Nicole Drazenovich, Janet E Foley, Richard N. Brown

Research output: Contribution to journalArticle

71 Citations (Scopus)

Abstract

Anaplasma phagocytophilum is an emerging pathogen throughout much of the Holarctic, where Ixodes spp. tick vectors occur. This organism was expected to be present at study sites in Humboldt County, north-western California, based on the presence of appropriate tick vectors, seropositivity in sentinel hosts, and previously reported human infections. However, despite high seroprevalence suggesting circulating A. phagocytophilum, active infections in dogs and wildlife (including suspected reservoir species) were rare using published polymerase chain reaction (PCR) protocols. This finding was possible if the published PCR protocol lacked sensitivity for strains in the study areas. We report a new TaqMan-PCR (TM-PCR) assay targeting the msp2 gene that has greater sensitivity and specificity for diverse A. phagocytophilum strains from this region. The new assay detected as few as one plasmid copy and a range of genetically diverse strains of A. phagocytophilum. Specificity was confirmed by failure to amplify targets of closely related bacteria. Application of the TM-PCR to samples from northern California confirmed PCR-positivity in 94 woodrats (71%; n = 134), three (4%; n = 80) bears, and seven (7%; n = 97) domestic dogs. The msp2 TM-PCR protocol appears to be more sensitive for use in assays of samples from parts of western North America and possibly in other regions where populations are genetically diverse or divergent from eastern United States strains of A. phagocytophilum.

Original languageEnglish (US)
Pages (from-to)83-90
Number of pages8
JournalVector-Borne and Zoonotic Diseases
Volume6
Issue number1
DOIs
StatePublished - Mar 2006

Fingerprint

Anaplasma phagocytophilum
Domestic Animals
Protein Transport
Real-Time Polymerase Chain Reaction
Polymerase Chain Reaction
Infection
Proteins
Ticks
Dogs
Ixodes
Gene Targeting
Seroepidemiologic Studies
North America
Plasmids
Bacteria
Sensitivity and Specificity
Population

Keywords

  • Canis familiaris
  • Granulocytic anaplasmosis
  • Neotoma fuscipes
  • TaqMan
  • Ursus americanum

ASJC Scopus subject areas

  • Infectious Diseases
  • Parasitology

Cite this

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abstract = "Anaplasma phagocytophilum is an emerging pathogen throughout much of the Holarctic, where Ixodes spp. tick vectors occur. This organism was expected to be present at study sites in Humboldt County, north-western California, based on the presence of appropriate tick vectors, seropositivity in sentinel hosts, and previously reported human infections. However, despite high seroprevalence suggesting circulating A. phagocytophilum, active infections in dogs and wildlife (including suspected reservoir species) were rare using published polymerase chain reaction (PCR) protocols. This finding was possible if the published PCR protocol lacked sensitivity for strains in the study areas. We report a new TaqMan-PCR (TM-PCR) assay targeting the msp2 gene that has greater sensitivity and specificity for diverse A. phagocytophilum strains from this region. The new assay detected as few as one plasmid copy and a range of genetically diverse strains of A. phagocytophilum. Specificity was confirmed by failure to amplify targets of closely related bacteria. Application of the TM-PCR to samples from northern California confirmed PCR-positivity in 94 woodrats (71{\%}; n = 134), three (4{\%}; n = 80) bears, and seven (7{\%}; n = 97) domestic dogs. The msp2 TM-PCR protocol appears to be more sensitive for use in assays of samples from parts of western North America and possibly in other regions where populations are genetically diverse or divergent from eastern United States strains of A. phagocytophilum.",
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