Image: Loopamp Realtime Turbidimeter for Loop-mediated Isothermal Amplification (Photo courtesy of Eiken Chemical).
Molecular methods have great potential for sensitive parasite detection in the diagnosis of human African trypanosomiasis (HAT), but the requirements in terms of laboratory infrastructure limit their use to reference centers.
A recently develop commercial assay that uses loop mediated isothermal amplification (LAMP) has been transformed into a ready-to-use kit format that does not require thermocyclers and amplification and can be conducted in a heating block or a hot water bath.
Tropical medicine specialists at the Institute of Tropical Medicine (Antwerp, Belgium) working with colleagues at the Kinshasa University (Kinshasa, Democratic Republic of the Congo) evaluated the diagnostic performance of the Loopamp Trypanosoma brucei
assay (LAMP) in confirmed T.b. gambiense
HAT patients, HAT suspects and healthy endemic controls from the Democratic Republic of the Congo (DRC).
Archived DNA from blood samples from 142 T. b. gambiense
HAT patients, 111 healthy endemic controls and 97 HAT suspects with unconfirmed status were included in a retrospective evaluation. The Loopamp Trypanosoma brucei
Detection Kit (Eiken Chemical; Tokyo, Japan) was applied in duplicate on the DNA extracts. LAMP amplified T. brucei
DNA was visualized using the provided ultraviolet-light emitted diode (UV-LED) device. Amplified DNA emits green fluorescence while there is no fluorescence in negative samples.
Of the 142 HAT patients, 132 and 124 were LAMP positive in respectively the first and second run, corresponding with sensitivities of 93.0% and 87.3%. Of the 11 patients with trypanosomes detected only in lymph or in cerebral spinal fluid (CSF), seven were positive in both LAMP runs on blood. Of the 97 HAT suspects, six were positive in both replicates of LAMP, eight and 20 were positive in the first and second replicate, respectively. Of 111 healthy endemic controls, four tested positive twice with LAMP and four tested positive only once.
The authors concluded that the LAMP has similar diagnostic accuracy as the 18S ribosomal DNA (rDNA) polymerase chain reaction (PCR) and can replace PCR for accurate and simplified detection of Trypanozoon DNA in clinical specimens. LAMP showed good diagnostic accuracy and excellent reproducibility, and may have an important role to play in disease surveillance. However, it should be noted that the specificity of LAMP is not 100%, that HAT treatment is complex and toxic, and that the positive predictive value of tests is low in hypoendemic settings. The study was published on October 17, 2013, in the journal Public Library of Science Neglected Tropical Diseases.
Institute of Tropical Medicine