A comparative analysis of microscopy and PCR-based detection methods for blood parasites

We compared information obtained by both microscopy and nested mitochondrial cytochrome b PCR in determining prevalence of haemosporidian infections in naturally infected birds. Blood samples from 472 birds of 11 species belonging to 7 families and 4 orders were collected in Europe, Africa and North America. Skilled investigators investigated them using the PCR-based screening and microscopic examination of stained blood films. The overall prevalence of haemosporidian infections, which was determined combining results of both these methods, was 60%. Both methods slightly underestimated the overall prevalence of infection, which was 54.2% after the PCR diagnostics and 53.6% after microscopic examination. Importantly, both these tools showed the same trends of prevalence of Haemoproteus spp. (21% by PCR and 22% by microscopy), Plasmodium spp. (17% and 22%) and Leucocytozoon spp. (30% and 25%) in the same sample, testifying that microscopy is a reliable tool in determining patterns of distribution of blood haemosporidian parasites in naturally infected birds. We encourage using optical microscopy in studies of blood parasites in parallel to the now widely employed molecular methods. Microscopy is relatively inexpensive and provides valuable information about directions how molecular methods can be further improved and most effectively applied, especially in the field studies of parasites. Importantly, blood films, which are used for microscopic examination, should be of good quality; they should be examined properly by skilled investigators. In spite of relatively long duration of microscopy of each sample, such examination provides opportunities for simultaneous determination and verification of taxonomically different parasites. Presently, different PCR protocols must be used for the detection of parasites belonging to different genera; this is expensive and time-consuming.