Development of a tick-borne pathogen QPCR panel for detection of Anaplasma, Ehrlichia, Rickettsia, and Lyme disease Borrelia in animals
Anaplasma spp., Ehrlichia spp., Rickettsia spp., and Lyme disease associated Borrelia spp. are the most common tick-borne pathogens reported to infect human beings worldwide and other animals, such as dogs and horses. In the present study, we developed a broad-coverage SYBR Green QPCR panel consisti...
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| Published in | Journal of microbiological methods Vol. 151; pp. 83 - 89 |
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| Main Authors | , , , , |
| Format | Journal Article |
| Language | English |
| Published |
Netherlands
Elsevier B.V
01.08.2018
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| Abstract | Anaplasma spp., Ehrlichia spp., Rickettsia spp., and Lyme disease associated Borrelia spp. are the most common tick-borne pathogens reported to infect human beings worldwide and other animals, such as dogs and horses. In the present study, we developed a broad-coverage SYBR Green QPCR panel consisting of four individual assays for the detection and partial differentiation of the aforementioned pathogens. All assays were optimized to the same thermocycling condition and had a detection limit of 10 copies per reaction. The assays remained sensitive when used to test canine and equine blood DNA samples spiked with known amounts of synthetic DNA (gBlock) control template. The assays were specific, as evidenced by lack of cross reaction to non-target gBlock or other pathogens commonly tested in veterinary diagnostic labs. With appropriate Ct cutoff values for positive samples and negative controls and the melting temperature (TM) ranges established in the present study, the QPCR panel is suitable for accurate, convenient and rapid screening and confirmation of tick-borne pathogens in animals.
•A SYBR Green QPCR panel for the detection of four tick-borne pathogens in canine and equine samples•Simultaneous detection of Anaplasma, Ehrlichia,Rickettsia, and Borrelia burgdorferi•Sensitivity: 10 copy gBlock per reaction for clinical samples•Specificity: no cross reaction to non-target gBlock or other pathogens |
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| AbstractList | Anaplasma spp., Ehrlichia spp., Rickettsia spp., and Lyme disease associated Borrelia spp. are the most common tick-borne pathogens reported to infect human beings worldwide and other animals, such as dogs and horses. In the present study, we developed a broad-coverage SYBR Green QPCR panel consisting of four individual assays for the detection and partial differentiation of the aforementioned pathogens. All assays were optimized to the same thermocycling condition and had a detection limit of 10 copies per reaction. The assays remained sensitive when used to test canine and equine blood DNA samples spiked with known amounts of synthetic DNA (gBlock) control template. The assays were specific, as evidenced by lack of cross reaction to non-target gBlock or other pathogens commonly tested in veterinary diagnostic labs. With appropriate Ct cutoff values for positive samples and negative controls and the melting temperature (TM) ranges established in the present study, the QPCR panel is suitable for accurate, convenient and rapid screening and confirmation of tick-borne pathogens in animals. Anaplasma spp., Ehrlichia spp., Rickettsia spp., and Lyme disease associated Borrelia spp. are the most common tick-borne pathogens reported to infect human beings worldwide and other animals, such as dogs and horses. In the present study, we developed a broad-coverage SYBR Green QPCR panel consisting of four individual assays for the detection and partial differentiation of the aforementioned pathogens. All assays were optimized to the same thermocycling condition and had a detection limit of 10 copies per reaction. The assays remained sensitive when used to test canine and equine blood DNA samples spiked with known amounts of synthetic DNA (gBlock) control template. The assays were specific, as evidenced by lack of cross reaction to non-target gBlock or other pathogens commonly tested in veterinary diagnostic labs. With appropriate Ct cutoff values for positive samples and negative controls and the melting temperature (TM) ranges established in the present study, the QPCR panel is suitable for accurate, convenient and rapid screening and confirmation of tick-borne pathogens in animals. •A SYBR Green QPCR panel for the detection of four tick-borne pathogens in canine and equine samples•Simultaneous detection of Anaplasma, Ehrlichia,Rickettsia, and Borrelia burgdorferi•Sensitivity: 10 copy gBlock per reaction for clinical samples•Specificity: no cross reaction to non-target gBlock or other pathogens |
| Author | Zhang, Michael Z. Mitchell, William J. Stich, Roger W. Zhang, Shuping Shen, Zhenyu |
| Author_xml | – sequence: 1 givenname: Zhenyu surname: Shen fullname: Shen, Zhenyu organization: Veterinary Medical Diagnostic Laboratory, College of Veterinary Medicine, University of Missouri, Columbia, MO, USA – sequence: 2 givenname: Michael Z. surname: Zhang fullname: Zhang, Michael Z. organization: Veterinary Medical Diagnostic Laboratory, College of Veterinary Medicine, University of Missouri, Columbia, MO, USA – sequence: 3 givenname: Roger W. surname: Stich fullname: Stich, Roger W. organization: Department of Veterinary Pathobiology, College of Veterinary Medicine, University of Missouri, Columbia, MO, USA – sequence: 4 givenname: William J. surname: Mitchell fullname: Mitchell, William J. organization: Veterinary Medical Diagnostic Laboratory, College of Veterinary Medicine, University of Missouri, Columbia, MO, USA – sequence: 5 givenname: Shuping surname: Zhang fullname: Zhang, Shuping email: zhangshup@missouri.edu organization: Veterinary Medical Diagnostic Laboratory, College of Veterinary Medicine, University of Missouri, Columbia, MO, USA |
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| CitedBy_id | crossref_primary_10_46909_alse_552059 crossref_primary_10_1186_s13071_021_04756_9 crossref_primary_10_1016_j_ttbdis_2020_101462 crossref_primary_10_1186_s12575_023_00226_x crossref_primary_10_1016_j_jevs_2019_05_003 crossref_primary_10_1590_0103_8478cr20200891 crossref_primary_10_14202_vetworld_2022_1699_1705 crossref_primary_10_3390_microorganisms9051092 crossref_primary_10_3390_pathogens10121541 |
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| Keywords | Tick-borne Anaplasma Borrelia Real-time PCR Rickettsia Ehrlichia |
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| SubjectTerms | Anaplasma Anaplasma - genetics Anaplasma - isolation & purification Anaplasma - pathogenicity Animals Borrelia Borrelia - genetics Borrelia - isolation & purification Borrelia - pathogenicity Ehrlichia Ehrlichia - genetics Ehrlichia - isolation & purification Ehrlichia - pathogenicity Horse Diseases - microbiology Horses Lyme Disease - diagnosis Lyme Disease - microbiology Molecular Diagnostic Techniques - methods Real-time PCR Real-Time Polymerase Chain Reaction - methods Rickettsia Rickettsia - genetics Rickettsia - isolation & purification Rickettsia - pathogenicity Sensitivity and Specificity Temperature Tick-borne Tick-Borne Diseases - diagnosis Tick-Borne Diseases - microbiology Tick-Borne Diseases - veterinary Ticks - microbiology |
| Title | Development of a tick-borne pathogen QPCR panel for detection of Anaplasma, Ehrlichia, Rickettsia, and Lyme disease Borrelia in animals |
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