title: Nested cytochrome b polymerase chain reaction diagnostics underestimate mixed infections of avian blood haemosporidian parasites: microscopy is still essential.
abstract: Numerous polymerase chain reaction (PCR)-based methods have been developed and used increasingly to screen vertebrate blood samples for the diagnosis of haemosporidian blood parasites (Sporozoa, Haemosporida), but a rigorous evaluation of the sensitivity of these methods for detecting mixed infections of different haemosporidian species belonging to the same and different genera and subgenera is lacking. This study links the information obtained by nested cytochrome b PCR and traditional microscopy in determining mixed haemosporidian infections in naturally infected birds. Samples from 83 individual passerine birds with single infections of Haemoproteus or Plasmodium spp., as determined by mitochondrial DNA amplification, also were investigated by microscopic examination of stained blood films. Thirty-six samples (43%) were found to harbor mixed Haemoproteus, or Plasmodium spp. infections, or both. Thus, the PCR assays alone underestimate the occurrence of mixed infections of haemosporidian parasites in naturally infected birds. To determine the true species composition of the haemosporidians in each individual host, PCR diagnostics need to be improved. Specific primers for Haemoproteus spp. and Plasmodium spp. should be developed. Ideally, a combination of the approaches of both microscopy and PCR-based methods is recommended for this purpose.
authors.affiliation: Institute of Ecology, Vilnius University, Vilnius 21, LT-08412, Lithuania. gedvalk@ekoi.lt
authors.forename: Gediminas
authors.initials: G
authors.lastname: Valkiŭnas
authors.forename: Staffan
authors.initials: S
authors.lastname: Bensch
authors.forename: Tatjana A
authors.initials: TA
authors.lastname: Iezhova
authors.forename: Asta
authors.initials: A
authors.lastname: Krizanauskiené
authors.forename: Olof
authors.initials: O
authors.lastname: Hellgren
authors.forename: Casimir V
authors.initials: CV
authors.lastname: Bolshakov
mesh_terms: D000818:Animals; D001715:Bird Diseases / Q000175:diagnosis* / Q000469:parasitology; D045303:Cytochromes b / Q000235:genetics*; D004272:DNA, Mitochondrial / Q000032:analysis; D016054:DNA, Protozoan / Q000032:analysis; D016787:Haemosporida / Q000201:enzymology / Q000235:genetics / Q000302:isolation & purification*; D046109:Passeriformes / Q000469:parasitology*; D016133:Polymerase Chain Reaction / Q000592:standards / Q000662:veterinary*; D011529:Protozoan Infections, Animal / Q000175:diagnosis* / Q000469:parasitology; D012680:Sensitivity and Specificity; D013045:Species Specificity
chemical_list: D004272:DNA, Mitochondrial; D016054:DNA, Protozoan; D045303:Cytochromes b