Rapid detection of active human cytomegalovirus infection in pregnancy using loop-mediated isothermal amplification

Abstract

Understanding the association between congenital human cytomegalovirus (HCMV) infection and active maternal HCMV infection during pregnancy is important for maternal and neonatal healthcare. In the present study, a loop-mediated isothermal amplification (LAMP) method was established for the detection of CMV DNA from whole blood or amniotic fluid samples, using reverse transcription-quantitative polymerase chain reaction. The results of the present study demonstrated that the CMV LAMP assay detection was specific for CMV DNA, whereas it did not detect viral DNA from herpes simplex type 1 (HSV-1), HSV-2, varicella zoster virus, HSV-6 or HSV-7. Sensitivity determination using serially-diluted CMV glycoprotein B-containing plasmids, demonstrated that >10 copies per tube were detectable using the CMV LAMP method. Furthermore, the detection results, using the LAMP method for 336 whole blood samples, demonstrated that at a threshold of 10(1)-10(4) copies per tube, the sensitivity of this method was 86.96-100%, the specificity was 97.24-100%, the positive predictive value was 76.92-100% and the negative predictive value was 99.05-100%. The results for 11 amniotic fluid samples from pregnant women with whole blood CMV-positive and 15 control amniotic fluid samples, indicated that the CMV LAMP assay was sensitive and specific for CMV detection. In conclusion, in the present study, a CMV LAMP method was developed, which was shown to be sensitive, specific and efficient in the detection of HCMV infection. Furthermore, CMV LAMP is capable of detecting active CMV infection in pregnant women. Therefore, the current study provides novel insights into diagnostic approaches for active CMV infection in pregnant women.

Development and application of loop-mediated isothermal amplification for detecting the highly benzimidazole-resistant isolates in Sclerotinia sclerotiorum

“…Since LAMP was reported in 2000 by Notomi, it has been widely applied in nucleic acid analysis because of its simplicity, rapidity, high efficiency, and outstanding specificity 12 . In recent years, this method has been successfully applied in molecular detection and diagnostics of bacterial 16 17 , viral 18 19 , fungal 20 21 22 23 , and parasitic diseases 24 25 in both animals and plants. However, the application of LAMP in fungicide resistance of plant pathogens has not been reported until the detection of benzimidazole-resistant mutants of F. graminearum was disclosed by our group 26 .…”

Comparison of Reverse Transcription Loop-Mediated Isothermal Amplification Method with SYBR Green Real-Time RT-PCR and Direct Fluorescent Antibody Test for Diagnosis of Rabies

“…However, Nagamine et al suggested that if circular primers are used, RT-LAMP reaction could be performed in 30 min (26). The study by Wang et al also identified CMV virus at 64 C in 30 min with RT-LAMP reaction (27).…”

Development and application of a simple, rapid and sensitive method for detecting moderately carbendazim‐resistant isolates in Botrytis cinerea

“…LAMP, a novel DNA amplification technique, is an auto‐cycling and strand displacement DNA synthesis method involving the use of the large fragment of Bst DNA polymerase and a set of 4–6 specific primers (Notomi et al, ; Mori et al, ; Nagamine et al, ; Curtis et al, ; Goto et al, ). Since this technique was developed in 2000, LAMP has been widely used for detection of pathogens of both plants and animals (Duan et al, ,b; Kogovšek et al, ; Lee et al, ; Niu et al, ; Okuda et al, ; Wang et al, ; Zhuo et al, ). However, it has been rarely used for detection of mutations conferring drug resistance in pathogenic microorganisms until recently.…”