Abstract
Ureaplasma urealyticum (UU) is commonly present in human reproductive tract, which frequently leads to genital tract infection. Hence, there is an urgent need to develop a rapid detection method for UU. In our study, a real-time fluorescence loop-mediated isothermal amplification (LAMP) assay was developed and evaluated for the detection of UU. Two primers were specifically designed based on the highly conserved regions of ureaseB genes. The reaction was carried out for 60 min in a constant temperature system using Bst DNA polymerase, and the process was monitored by real-time fluorescence signal, while polymerase chain reaction (PCR) was performed simultaneously. In real-time fluorescence LAMP reaction system, positive result was only obtained for UU among 9 bacterial strains, with detection sensitivity of 42 pg/μL (4.2 × 105 CFU/mL), and all 16 clinical samples of UU could be detected. In conclusion, real-time fluorescence LAMP is a simple, sensitive, specific and effective method compared with conventional PCR, which shows great promise in the rapid detection of UU.
Highlights
Reproductive tract infection caused by Mycoplasma hominis (MH), Mycoplasma genitalium, and Ureaplasma urealyticum (UU), imposes a threat to human reproductive tract health
Our study aims to evaluate real-time fluorescence loop-mediated isothermal amplification (LAMP) assay with a modification, which combined with high sensitivity, specificity and efficiency, to investigate its feasibility in clinical practice and facilitate the rapid detection of UU in clinical samples
Specificity of real‐time fluorescence LAMP In the present study, eight common pathogenic bacteria were selected as negative controls and no amplification was tested by real-time fluorescence LAMP assay (Fig. 2)
Summary
Reproductive tract infection caused by Mycoplasma hominis (MH), Mycoplasma genitalium, and Ureaplasma urealyticum (UU), imposes a threat to human reproductive tract health. UU is the most prevalent pathogen that colonizes the human reproductive tract (Esen et al 2017). UU causes severe problems for infants and their mothers. Conventional diagnosis of UU infection relies on positive culture, antigen detection, serology, and molecular biology. Polymerase chain reaction (PCR) is one of the most common techniques (Liu et al 2017). The above-mentioned methods require either long culture periods or precision instruments, limiting their application in primary medical institutions, especially in developing countries. Apart from that, PCR increases the risk of sample cross-contamination (Dehghan Esmatabadi et al 2015). An alternative diagnostic is needed to diagnose UU infection fast and efficiently
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