Challenges in the Diagnostic Performance of Parasitological and Molecular Tests in the Detection of African Trypanosomiasis in Cattle in Mambwe District in Eastern Zambia

Abstract Background: Trypanosomiasis is a Neglected Tropical Disease with serious health and economic implications. Disease eradication and control programs rely on active case detection through mass population screening. Screening tools and techniques therefore need to be adequately sensitive, practically quick to perform, and affordable. This study compared the field application of the polymerase chain reaction, the loop mediated isothermal amplification technique, and microscopy, in the detection of trypanosomes in cattle blood in Mambwe district in eastern Zambia. Methods: Blood samples were collected from 227 cattle into three heparinised micro capillary tubes, and tested for trypanosomiasis infection using microscopy, ITS-PCR and RIME-LAMP. The comparative diagnostic performance of each of the methods was evaluated using the chi-square test, kappa test and receive operator curves. Results: Microscopy on buffy coat detected 17 cases (n=227), by thin smears detected 26 cases (n=227), and by thick smears detected 28 cases (n=227). In total, microscopy detected 40 cases (n=227). ITS-PCR- on blood spots stored on filter paper detected 47 cases (n=227), ITS-PCR- on blood spots stored on FTA cards detected 83 cases (n=227) and RIME-LAMP-FTA detected 18 cases (n = 131). Using microscopy as gold standard, sensitivity and specificity of ITS-PCR was compared. ITS-PCR-FTA had a better specificity and sensitivity (SE=77.5%; SP=72.2%; k = 0.35) than ITS-PCR-FP (SP = 88%; SE = 60%; kappa = 0.45). Prevalence of Trypanosoma brucei s.l. was higher on RIME-LAMP-FTA (18/131) than ITS-PCR-FTA (19/227). Conclusion: Our results are not perfect but are a good illustration of the current diagnostic challenges in rural Africa. Findings showed that none of the diagnostic tests could be taken as having performed better than the others and that each of the tests offered some advantages and limitations. In endemic rural areas of Africa, the use of PCR and LAMP requires specialised staff, laboratory supplies and infrastructure which is often not available. For this reason, microscopy remains the most practical option for field diagnosis of trypanosomes but understanding its limitations is critical particularly when applied for surveillance purposes.