Anaplasma platys: an improved PCR for its detection in dogs

• Published in: Experimental parasitology Vol. 109; no. 3; pp. 176 - 180
• Main Authors: Martin, Anthony R., Brown, Graeme K., Hugh Dunstan, R., Roberts, Timothy K.
• Format: Journal Article
• Language: English
• Published: San Diego, CA Elsevier Inc 01.03.2005; Elsevier
• Subjects: Anaplasma; Anaplasma - genetics; Anaplasma - isolation & purification; Anaplasma platys; anaplasmosis; Anaplasmosis - blood; Anaplasmosis - diagnosis; Anaplasmosis - parasitology; Animals; Biological and medical sciences; blood; Blood Platelets - parasitology; detection; disease diagnosis; DNA primers; DNA Primers - chemistry; DNA, Bacterial - blood; Dog Diseases - blood; Dog Diseases - diagnosis; Dog Diseases - parasitology; Dogs; Ehrlichia; Ehrlichia - genetics; Ehrlichia - isolation & purification; Fundamental and applied biological sciences. Psychology; Life cycle. Host-agent relationship. Pathogenesis; Nested PCR; nested polymerase chain reaction; parasitemia; PCR; Polymerase chain reaction; Polymerase Chain Reaction - veterinary; Protozoa; Reproducibility of Results; ribosomal DNA; RNA, Bacterial - genetics; RNA, Ribosomal, 16S - genetics; Sensitivity and Specificity; Australia; Polymerase chain reaction; Nested PCR; Anaplasma platys; Ehrlichia; PCR; Ehrlichiaceae; Fissipedia; Carnivora; Anaplasmataceae; Parasite; Ehrlichieae; Anaplasma; Canis familiaris; Rickettsiales; PCR: Polymerase chain reaction; Vertebrata; Mammalia; Anaplasma plutys; Bacteria; Detection; Nest; Dog
• ISSN: 0014-4894; 1090-2449
• DOI: 10.1016/j.exppara.2004.11.007

This study compares two PCR assays for the detection of Anaplasma platys in dog blood using primers based on the A. platys 16S rRNA gene. The first approach utilized a “standard” PCR protocol composed of a “single-step” direct amplification using an Ehrlichia genus-specific primer set. The second assay being a “nested” PCR screen that first involved a universal bacterial primer set that amplified the majority of the 16S rRNA gene, followed by the nested round of PCR using an A. platys-specific primer set. Of the 22 dogs sampled, 10 were found to contain A. platys DNA using both protocols, and an additional two dogs were found positive using the nested technique. An extract of A. platys positive genomic DNA was serially diluted and comparison of sensitivities determined between the nested PCR, and a direct assay using A. platys-specific primers. The nested protocol demonstrated an increased sensitivity by at least 2 orders of magnitude when compared to the direct assay alone. Our results indicated that the nested PCR assay with its increased sensitivity would be useful for experimental research investigations as well as offer the potential for use as a routine test in diagnostic pathology.