A nested PCR for the ssrRNA gene detects Trypanosoma binneyi in the platypus and Trypanosoma sp. in wombats and kangaroos in Australia

• Published in: International journal for parasitology Vol. 29; no. 2; pp. 331 - 339
• Main Authors: NOYES, H. A, STEVENS, J. R, TEIXEIRA, M, PHELAN, J, HOLZ, P
• Format: Journal Article
• Language: English
• Published: Oxford Elsevier Science 01-02-1999
• Subjects: Animal protozoal diseases; Animals; Animals, Wild; Australia; Biological and medical sciences; DNA, Protozoan - analysis; DNA, Ribosomal - analysis; Genes, rRNA; Infectious diseases; Macropodidae - parasitology; Macropus giganteus; Medical sciences; Molecular Sequence Data; Ornithorhynchus anatinus; Parasitic diseases; Platypus - parasitology; Polymerase Chain Reaction - methods; Polymorphism, Restriction Fragment Length; Protozoal diseases; Sequence Analysis, DNA; Trypanosoma; Trypanosoma - classification; Trypanosoma - genetics; Trypanosoma - isolation & purification; Trypanosomiasis - parasitology; Trypanosomiasis - veterinary; Vombatus ursinus; Australia; Kinetoplastida; Protozoa; Host parasite relation; Protozoal disease; Methodology; Ornithorhynchus anatinus; Macropus giganteus; Parasite; Host; Zoopathogen; Parasitosis; Method; Monotremata; Subunit; Trypanosoma cruzi; Infection; Polymerase chain reaction; Vertebrata; Mammalia; Restriction fragment length polymorphism; Ribosomal RNA; Marsupialia; Detection; Trypanosomiasis
• ISSN: 0020-7519; 1879-0135
• DOI: 10.1016/S0020-7519(98)00167-2

Trypanosome infections in their natural hosts are frequently difficult to detect by microscopy, and culture methods are unreliable and not suitable for all species of Trypanosoma. A nested PCR strategy for detecting and identifying Trypanosoma species, suitable for detecting both known and unknown trypanosomes, is presented. Thirty-two blood samples from 23 species of Australian birds and mammals were screened by a nested PCR for the presence of Trypanosoma sp. ssrRNA. Three infections were detected, one in an eastern grey kangaroo (Macropus giganteus), one in a common wombat (Vombatus ursinus) and one in a platypus (Ornithorhynchus anatinus). The kangaroo and wombat are new host records for Trypanosoma sp.; the platypus parasite was Trypanosoma hinneyi. The three parasites could be distinguished by restriction fragment length polymorphisms of the amplified fragment of the ssrRNA gene. The kangaroo and wombat parasites were also isolated in a semi-solid blood agar medium. The culture forms of the kangaroo trypanosome had an expanded flagellar sheath in which structures similar to hemidesmosomes were detected by EM. The nested PCR was at least as sensitive as culture, and analysis of the PCR products gave parasite-specific fingerprints. Therefore this method could be suitable for rapidly screening host animals for the presence of trypanosomes and identifying the infecting strain.