Application and development of molecular diagnostic techniques in human assisted reproduction / 中华检验医学杂志

In the past few years, with the breakthrough development of DNA sequencing technology, preimplantation genetic testing (PGT) has been widely used as an important part of assisted reproductive technology (ART). The progress of next generation sequencing (NGS) technology and the improvement of resolution, as well as the comprehensive application of molecular diagnostic technology makes it possible to perform the extensive and comprehensive chromosome screening and the clinical valuable detection of small gene fragment missing and repetition simultaneously, which is of great significance in terms of improving the pregnancy rate and reducing the abortion rate, multiplets rate and malformation rate. The common PGT molecular diagnostic techniques involves fluorescence in situ hybridization (FISH), single nucleotide polymorphism array(SNP-array), quantitative polymerase chain reaction(qPCR) and NGS. And each of them has its own highlights in clinical application and there are many uncertainties that are difficult to control. Moreover, ethical concerns brought about by technological progress also need to be addressed.

Application and development of molecular diagnostic techniques for detection infectious diseases / 中华检验医学杂志

With the development of the concept of precision medicine, under the background of the new coronavirus pneumonia epidemic, the clinical diagnosis and treatment of infectious diseases has received more and more attention. The experimental diagnosis technology with molecular biology as the core is used as important means for the clinical laboratory diagnosis of infectious diseases. This lcind of technology is paid special attention. In recent years, advances in nanomaterials, applied chemistry, photophysics, and biosensing technologies have also ushered in revolutionary and creative developments in molecular diagnostic technology. This article reviews the application and development of the latest molecular diagnostic technologies, such as next-generation quantitative PCR technology and gene sequencing technology, isothermal amplification technology, biochip and biosensor technology in the clinical diagnosis of infectious diseases.

New molecular diagnostic technologies for clinical detection of SARS-CoV-2.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused an ongoing pandemic of new coronavirus pneumonia (corona virus disease 2019, COVID-19). The virus has a long incubation period and strong infectivity, which poses a major threat to global health and safety. Detection of SARS-CoV-2 nucleic acid lies at the center of rapid detection of COVID-19, which is instrumental for mitigation of the ongoing pandemic. As of August 17, 2020, The National Medical Products Administration in China has approved 15 new coronavirus nucleic acid detection kits, 10 kits of which are based on reverse transcription-real-time quantitative PCR (RT-qPCR) technology. The remaining kits use five molecular diagnostic technologies different from RT-qPCR. This article reviews the principles, reaction time, advantages and disadvantages of above 15 detection kits, in order to provide references for rapid screening, diagnosis, prevention and control of COVID-19 and similar infectious diseases.

A colorimetric Loop-mediated isothermal amplification (LAMP) assay based on HRP-mimicking molecular beacon for the rapid detection of Vibrio parahaemolyticus.

In the world wide, food poisoning accidents related to Vibrio spp. are on the rise, even numbers of food poisoning by other foodborne pathogens are decreasing. Therefore, the requirement of the rapid, sensitive and convenient detection method for V. parahaemolyticus has been grown. The objective of this study is to develop a colorimetric loop-mediated isothermal amplification (LAMP) assay using a molecular beacon (HRPzyme connected with complementary oligonucleotides at the 5' and 3' ends) for the rapid, sensitive, and convenient detection of V. parahaemolyticus. The colorimetric LAMP assay optimized at 58.8°C showed a detection limit of 1 × 100 CFU/mL and was confirmed to be specific to V. parahaemolyticus. The colorimetric LAMP assay can be finished within 1 h including DNA extraction step. The method was successfully applied to flatfish samples artificially inoculated with known amount of V. parahaemolyticus, and its cut-off value for the flatfish samples was 1 × 101 CFU/g. In addition, the colorimetric LAMP assay developed in the study was found to be able to correct false-positive results, which are known to be a disadvantage of conventional LAMP assays. Therefore, these results indicated that the colorimetric LAMP method is a useful tool for the rapid, sensitive and convenient detection of V. parahaemolyticus in fishes and can also be used as a point-of-care molecular diagnostic technique since it does not require any expensive equipment such as a thermocycler and detectors.

Diagnóstico molecular de candidemia mediante PCR semianidada en pacientes críticos / Molecular diagnosis of candidemia by seminested PCR in critically ill patients

Objetivos: estandarizar una prueba molecular basada en PCR semianidada para el diagnóstico de candidemia Metodología: se desarrolló un estudio piloto en que se optimizó el protocolo de extracción y utilización de ADN ribosomal, amplificando la región ITS-2 completa de Candida spp. Y mediante PCR semianidada, reamplificando segmentos específicos de nueve especies clínicamente relevantes de Candida. Se utilizaron muestras clínicas de plasma de pacientes críticamente enfermos con candidemia documentada microbiológicamente del Hospital de la Samaritana en Bogotá. Se utilizó una muestra de candidemia simulada utilizando una cepa de Candida obtenida de orina. Resultados: se logró protocolizar dicha técnica y tener a partir de muestras clínicas de plasma una sensibilidad probable de 90% y especificidad teórica cercana a 100%. La capacidad de detección de este ensayo es hasta de una célula por 200 μl de plasma. Conclusiones: se estandarizó una prueba molecular para detección de candidemia en pacientes críticamente enfermos (Acta Med Colomb 2011; 36: 135-140).

Objectives: to standardize a molecular test based on semi-nested PCR for diagnosis of candidemia. Methods: we developed a pilot study by means of optimizing the extraction protocol and use of ribosomal DNA, amplifying the complete ITS-2 region of Candida spp. And by semi-nested PCR reamplifying specific segments of nine clinically relevant Candida species. Clinical samples of plasma of critically ill patients with microbiologically proven candidemia from a third level hospital in Bogotá. Another sample of simulated candidemia was used with Candida strain form a urine sample. Results: we were able to standardize this technique and achieved from clinical samples of plasma a likely sensitivity of 90% and a theoretical specificity of 100%. The detection capability of this test is up to 1 cell per 200 μl of plasma. Conclusion: a molecular test for detection of candidemia among critically ill patients was standardized (Acta Med Colomb 2011; 36: 135-140).