DNA Microarray Detects 18 Important Human Blood Protozoan Species

Accurate detection of blood protozoa from clinical samples is important for diagnosis, treatment and control of related diseases and developing a rapid, simple, and convenient detection method for protozoan detection is an urgent need.

Blood protozoa are single-cell organisms that often have flagella, cilia or other structures that help them move. They sometimes form parasitic relationships with humans and cause diseases or infections. The most common blood protozoa in humans, animals, and vectors include Plasmodium, Leishmania, Trypanosoma, Toxoplasma gondii and Babesia.

Scientists at Fudan University (Shanghai, PR China) and their colleagues collected a total of 438 samples from humans, animals and vectors from August 2012 to December 2014 as well as 100 blood samples from healthy individuals. Reference blood protozoan samples were either stored in the laboratory or kindly provided by different partner laboratories. Blood smears were prepared for each clinical blood sample and were examined under an oil immersion lens at a total magnification of ×1000 for presence of blood protozoa.

The whole blood that was suspected infected Leishmania from human and canine was also tested by the Kalazar Detect dipstick, a recombinant k39 antigen-based immunochromatographic strip, to detect anti-leishmanial antibodies (InBios International, Seattle, WA, USA). The team performed microarray assays simultaneously that identified 18 species of common blood protozoa based on the differences in respective target genes. A total of 20 specific primer pairs and 107 microarray probes were selected according to conserved regions, which were designed to identify 18 species in five blood protozoan genera.

The scientists found that the positive detection rate of the microarray assay was 91.8% (402/438). Sensitivity and specificity for blood protozoan detection ranged from 82.4% to 100.0% and 95.1% to 100.0%, respectively. Positive predictive value (PPV) and negative predictive value (NPV) ranged from 20.0% to 100.0% and 96.8% to 100.0%, respectively. Youden index varied from 0.82 to 0.98. The detection limit of the DNA microarrays ranged from 200 to 500 copies/reaction, which was similar to the polymerase chain reaction (PCR) findings. The concordance rate between microarray data and DNA sequencing results was 100%.

The authors concluded that overall, the microarray platform can provide a convenient, accurate and reliable diagnostic tool for the identification of 18 of the most common blood protozoan species. The system should be widely used in future detection assay and management of blood protozoan infections in busy hospitals and research institutes, and would help in disease control and prevention plans. The study was published on December 2, 2016, in the Public Library of Science Neglected Tropical Diseases.

Related Links:
Fudan University
InBios International