Field implementation of a 3D printer based DNA extraction method coupled to LAMP for congenital Chagas disease diagnosis

Abstract

Congenital Chagas disease entails the transmission of Trypanosoma cruzi infection from a mother to her child. With currently available chemotherapies, the cure rate for infected children is almost 100 % if administered early upon infection. It is therefore of great relevance to diagnose newborns on time. However, the algorithm to detect congenital T. cruzi infection involves the performance of microhematocrite or micromethod at delivery or during the first months of life and a confirmatory serology at 10 months of age. In highly endemic areas where people live far away from reference centers, many infants never go back to confirm the diagnosis and receive treatment if infected. The challenge is then to implement sensitive and rapid diagnostic techniques that can be performed in minimally equipped laboratories. At present there is a prototype loop isothermal amplification molecular test available (T. cruzi-LAMP kit, Eiken, Japan), with similar sensitivity to that of real time PCR (qPCR), but easier to use. Nonetheless, highly purified DNA is needed and obtaining it is time consuming and requires equipment unavailable in endemic regions. Thus, our aim was to couple the T. cruzi-LAMP kit to a recently developed DNA extraction device based on a low cost 3D printer (named PrintrLab), and to test its use in a hospital located in the “Gran Chaco”, a highly endemic region for Chagas disease. The PrintrLab was programmed to purify DNA from whole blood-EDTA samples and to provide the incubation step for the T. cruzi-LAMP reaction. The process took about 2.5 hours to yield a result, while manual DNA extraction and subsequent qPCR normally take more than 6. Performance of the “PrintrLab-LAMP” duo was tested with blood-EDTA samples artificially contaminated with 0, 1, 2, 5, 10 and 100 parasites eq/mL and a sensitivity around 2 pararasites eq/mL was achieved. Finally, 70 clinical samples from infants born to seropositive mothers were evaluated and all the micromethod positive ones, 6 samples in total, were detected by the "PrintrLab-LAMP" approach. In conclusion, the “PrintrLabLAMP” device showed a good sensitivity, the protocol was faster than other molecular techniques and it could be successfully used in a minimally equipped laboratory.