Ten-minute direct detection of Zika virus in serum samples by RT-LAMP

Abstract

Zika virus (ZIKV) is a current threat to global health. In most of cases, ZIKV infection has no symptoms; however in some cases, ZIKV can cause paralysis (Guillain-Barré syndrome), and in pregnant women, it can cause birth defects in infants. Rapid and accurate diagnosis can help improve disease control as well as being vital to prenatal care for women living in endemic areas. Molecular diagnostics based on isothermal amplification techniques are an excellent alternative to conventional methods of DNA amplification, such as PCR. Here, we develop and optimized a rapid and sensitive method for direct detection of ZIKV in Serum samples based on RT-LAMP and visual detection. The reaction was thermally controlled with a thermoblock for 10 min at 72 °C. The results show that the use of the Bst 3.0 enzyme and an adequate optimization can further reduce the time needed for the RT-LAMP reaction to detect ZIKV. Our results demonstrate that it is possible to detect ZIKV through RT-LAMP directly from a Serum sample, without prior RNA extraction. As little as 10^-3 copies of RNA in a 10 μL reaction (20 zepto-molar) was detected by RT-LAMP from a panel of 51 Serum samples (16 samples from pregnant women and 35 samples from newborns infected with ZIKV during pregnancy). The RT-LAMP has proven to be a valuable tool for molecular diagnosis of Zika, presenting a great potential for point-of-care applications, especially in developing countries.

Keywords: Molecular diagnosis; Point-of-care; RT-LAMP; Rapid detection; Zika virus.