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Beatriz Jorge Coelho, Bruno Veigas, Hugo Águas, Elvira Fortunato, Rodrigo Martins, Pedro Viana Baptista, Rui Igreja
Digital microfluidics (DMF) arises as the next step in the fast-evolving field of operation platforms for molecular diagnostics. Moreover, isothermal schemes, such as loop-mediated isothermal amplification (LAMP), allow for further simplification of amplification protocols. Integrating DMF with LAMP will be at the core of a new generation of detection devices for effective molecular diagnostics at point-of-care (POC), providing simple, fast, and automated nucleic acid amplification with exceptional integration capabilities...
November 16, 2017: Sensors
Jun Sakai, Takuya Maeda, Norihito Tarumoto, Kazuhisa Misawa, Shinsuke Tamura, Kazuo Imai, Toshiyuki Yamaguchi, Satoshi Iwata, Takashi Murakami, Shigefumi Maesaki
No abstract text is available yet for this article.
November 11, 2017: Journal of Microbiological Methods
Yi Wang, Dongxin Liu, Jianping Deng, Yan Wang, Jianguo Xu, Changyun Ye
Loop-mediated isothermal amplification (LAMP) is the most popular technique to amplify nucleic acid sequence without the use of temperature cycling. However, LAMP is often confounded by false-positive results, arising from interactions between (hetero-dimer) or within (self-dimerization) primers, off-target hybrids and carryover contaminants. Here, we devised a new LAMP technique that is self-avoiding molecular recognition system (SAMRS) components and antarctic thermal sensitive uracil-DNA-glycosylase (AUDG) enzyme-assisted, termed AUDG-SAMRS-LAMP...
December 15, 2017: Analytica Chimica Acta
Binan Zhao, Dan Yang, Yongchun Zhang, Yan Xu, Xiao Zhao, Jieling Liang, Xiaorui Fan, Yanan Du, Ziping Zhu, Bin Shi, Qi Zhang, Xiaoxia Zhang, Youming Cai, Kai Zhao
Lily mottle virus (LMoV; genus Potyvirus, family Potyviridae) infects plants of the genus Lilium, causing a reduction in flower and bulb quality. A rapid and sensitive loop-mediated isothermal amplification (LAMP) assay was developed to detect the coat protein gene of LMoV. This LAMP method was highly specific for LMoV, with no cross-reaction with other lily viruses. The sensitivity of LMoV using the LAMP assay was 100 times more sensitive than that using conventional polymerase chain reaction. A reverse transcription LAMP (RT-LAMP) was then successfully applied to detect LMoV RNA...
November 14, 2017: Archives of Virology
M A Diallo, K Diongue, G Diagne, M C Seck, M Ndiaye, B Dièye, Y D Ndiaye, A S Badiane, D Ndiaye
Recently in Senegal, a case of Plasmodium ovale malaria had led to a diagnostic difficulty due to the ignorance of this parasite and the neglect of it. The objective of this study was to actively investigate cases of P. ovale malaria that would be misdiagnosed in the health centre structures of Senegal. The study was conducted in three areas that reflect different epidemiological strata of malaria. Microscopy was performed by microscopy experts on suspected malaria patients. The results were validated by Rougemont real-time PCR...
November 10, 2017: Bulletin de la Société de Pathologie Exotique
Fábio Ferreira Carlos, Bruno Veigas, Ana S Matias, Gonçalo Doria, Orfeu Flores, Pedro V Baptista
Due to their relevance as disease biomarkers and for diagnostics, screening of single nucleotide polymorphism (SNPs) requires simple and straightforward strategies capable to provide results in medium throughput settings. Suitable approaches relying on isothermal amplification techniques have been evolving to substitute the cumbersome and highly specialized PCR amplification detection schemes. Nonetheless, identification of an individual's genotype still requires sophisticated equipment and laborious methods...
December 2017: Biotechnology Reports
Kelly J Domesle, Qianru Yang, Thomas S Hammack, Beilei Ge
Loop-mediated isothermal amplification (LAMP) has emerged as a promising alternative to PCR for pathogen detection in food testing and clinical diagnostics. This study aimed to validate a Salmonella LAMP method run on both turbidimetry (LAMP I) and fluorescence (LAMP II) platforms in representative animal food commodities. The U.S. Food and Drug Administration (FDA)'s culture-based Bacteriological Analytical Manual (BAM) method was used as the reference method and a real-time quantitative PCR (qPCR) assay was also performed...
October 18, 2017: International Journal of Food Microbiology
Krzysztof Treder, Joanna Chołuj, Bogumiła Zacharzewska, Lavanya Babujee, Mateusz Mielczarek, Adam Burzyński, Aurélie M Rakotondrafara
Potato virus Y (PVY) infection has been a global challenge for potato production and the leading cause of downgrading and rejection of seed crops for certification. Accurate and timely diagnosis is a key for effective disease control. Here, we have optimized a reverse transcription loop-mediated amplification (RT-LAMP) assay to differentiate the PVY O and N serotypes. The RT-LAMP assay is based on isothermal autocyclic strand displacement during DNA synthesis. The high specificity of this method relies heavily on the primer sets designed for the amplification of the targeted regions...
November 8, 2017: Archives of Virology
Liang Wan, Tianlan Chen, Jie Gao, Cheng Dong, Ada Hang-Heng Wong, Yanwei Jia, Pui-In Mak, Chu-Xia Deng, Rui P Martins
A digital microfluidic (DMF) system has been developed for loop-mediated isothermal amplification (LAMP)-based pathogen nucleic acid detection using specific low melting temperature (Tm) Molecular Beacon DNA probes. A positive-temperature-coefficient heater with a temperature sensor for real-time thermal regulation was integrated into the control unit, which generated actuation signals for droplet manipulation. To enhance the specificity of the LAMP reaction, low-Tm Molecular Beacon probes were designed within the single-stranded loop structures on the LAMP reaction products...
November 6, 2017: Scientific Reports
Rui Mao, Ge Ge, Zhuo Wang, Rongzhang Hao, Guohao Zhang, Zhenzhou Yang, Bincheng Lin, Yajun Ma, Hongtao Liu, Yuguang Du
Malaria infection poses a great threaten to public health even nowadays. The conventional diagnosis tools of malaria parasites and vectors require systematic training for the observers accompanied by the low throughput. In this study, a new detection system, i.e., multiplex microfluidic loop-mediated isothermal amplification (mμLAMP) array, was developed to provide a convenient, rapid and economical detection system for malaria diagnosis. A microfluidic-based detection chip was designed and developed, targeting the conserved gene of four Anopheles and two Plasmodium species responsible for most of the malaria cases occurred in China...
November 2, 2017: Acta Tropica
Kazuhiro Kawai, Mika Inada, Keiko Ito, Koji Hashimoto, Masaru Nikaido, Eiji Hata, Ken Katsuda, Yoshio Kiku, Yuichi Tagawa, Tomohito Hayashi
Bovine mastitis causes significant economic losses in the dairy industry. Effective prevention of bovine mastitis requires an understanding of the infection status of a pathogenic microorganism in a herd that has not yet shown clinical signs of mastitis and appropriate treatment specific for the pathogenic microorganism. However, bacterial identification by culture has drawbacks in that the sensitivity may be low and the procedure can be complex. In this study, we developed a genetic detection method to identify mastitis pathogens using a simple and highly sensitive electrochemical DNA chip which can specifically detect bacterial DNA in milk specimens...
November 1, 2017: Journal of Veterinary Medical Science
A Centeno-Cuadros, J L Tella, M Delibes, P Edelaar, M Carrete
PCR is a universal tool for the multiplication of specific DNA sequences. For example, PCR-based sex determination is widely used, and a diversity of primer sets is available. However, this protocol requires thermal cycling and electrophoresis so results are typically obtained in laboratories and several days after sampling. Loop-mediated isothermal amplification (LAMP) is an alternative to PCR that can take molecular ecology outside the laboratory. Although its application has been successfully probed for sex determination in three species of a single avian Family (Raptors, Accipitridae), its generality remains untested and suitable primers across taxa are lacking...
November 1, 2017: Molecular Ecology Resources
Sharmili Roy, Noor Faizah Mohd-Naim, Mohammadali Safavieh, Minhaz Uddin Ahmed
Nucleic acid detection is of paramount importance in monitoring of microbial pathogens in food safety and infectious disease diagnostic applications. To address these challenges, a rapid, cost-effective label-free technique for nucleic acid detection with minimal instrumentations is highly desired. Here, we present paper microchip to detect and quantify nucleic acid using colorimetric sensing modality. The extracted DNA from food samples of meat as well as microbial pathogens was amplified utilizing loop-mediated isothermal amplification (LAMP)...
November 10, 2017: ACS Sensors
Shigeru Onari, Takashige Okada, Takafumi Okada, Syuko Okano, Osamu Kakuta, Hirokazu Kutsuma, Masaaki Kobayashi, Yasuo Kondo, Naoya Sakaguchi, Takeshi Tajima, Masayoshi Nagao, Eiichi Nakayama, Ryo Niimi, Syuichi Nishimura, Yoshihito Higashidate, Toshiyuki Hikita, Hidenori Meguro, Toshihiko Mori, Yuko Yoto, Hiroyuki Tsutsumi
The sensitivity and specificity of a new rapid Mycoplasma pneumoniae antigen immunochromatography (IC) test, DK-MP-001, were determined using particle agglutination (PA) antibody response and loop-mediated isothermal amplification (LAMP) gene detection as the gold standard. Of 165 patients, 59 were diagnosed with M. pneumoniae infection based on a ≥fourfold rise of serum PA antibody during the course of the illness. Of the first visit swabs, 60 were positive for M. pneumoniae on LAMP, and 49 were positive for M...
October 2017: Pediatrics International: Official Journal of the Japan Pediatric Society
Koji Hashimoto, Mika Inada, Keiko Ito
We have developed a novel voltammetric DNA chip for real-time electrochemical detection of targeted nucleic acid sequences using loop-mediated isothermal amplification (LAMP) and ruthenium hexaamine (RuHex) as the intercalative redox compound. A GspSSD DNA polymerase was used for LAMP owing to its tolerance of the intercalative redox compound. The electrochemical reaction of 1 mM RuHex in the LAMP solution was measured continuously by linear sweep voltammetry at 65 °C using an electrochemical DNA chip. According to the LAMP reaction of the positive sample, the cathodic peak current of RuHex increased and the cathodic peak potential of RuHex shifted to negative voltage...
October 24, 2017: Analytical Biochemistry
X R Hu, D J Dai, H D Wang, C Q Zhang
Botrytis cinerea, a typical "high-risk" pathogenic fungus that rapidly develops resistance to fungicides, affects more than 1,000 species of 586 plant genera native to most continents and causes great economic losses. Therefore, a rapid and sensitive assay of fungicide resistance development in B. cinerea populations is crucial for scientific management. In this study, we established a Loop-mediated isothermal amplification (LAMP) system for the monitoring and evaluation of the risk of development of B. cinerea resistance to QoI fungicides; the method uses two LAMP assays...
October 24, 2017: Scientific Reports
Kai Feng, Wei Li, Zhihong Guo, Hong Duo, Yong Fu, Xiuying Shen, Cheng Tie, Rijie E, Changqin Xiao, Yanhong Luo, Guo Qi, Ma Ni, Qingmei Ma, Wataru Yamazaki, Ayako Yoshida, Yoichiro Horii, Kinpei Yagi, Nariaki Nonaka
For field-identification of taeniid cestodes in canine animals in Tibetan area, loop-mediated isothermal amplification (LAMP) assays for Echinococcus multilocularis, E. shiquicus, Taenia hydatigena, T. multiceps, T. pisiformis and T. crassiceps were developed and evaluated along with the reported assay for E. granulosus. The LAMP assays showed specific reaction with their corresponding target species DNA with the detection limit of 1 to 10 pg. Moreover, the assays for E. granulosus, E. multilocularis, T. hydatigena and T...
October 23, 2017: Journal of Veterinary Medical Science
Hua-Wei Chen, Wei-Mei Ching
Coxiella burnetii, the causative pathogen for Q fever, is an obligate intracellular bacterium and designated as a biosafety level 3 agent. Detection and quantification of the bacteria with conventional culturing methods is time-consuming and poses significant health risks. Polymerase chain reaction (PCR)-based assays have been developed for detecting C. burnetii and could provide rapid diagnosis. However, they require specialized equipment, including a cold chain for PCR reagents that maintains their stability during storage and transport...
October 2017: Heliyon
Mehran Khan, Benjin Li, Yue Jiang, Qiyong Weng, Qinghe Chen
Late blight, caused by the oomycete Phytophthora infestans, is one of the most devastating diseases affecting potato and tomato worldwide. Early diagnosis of the P. infestans pathogen causing late blight should be the top priority for addressing disease epidemics and management. In this study, we performed a loop-mediated isothermal amplification (LAMP) assay, conventional polymerase chain reaction (PCR), nested PCR, and real-time PCR to verify and compare the sensitivity and specificity of the reaction based on the Ypt1 (Ras-related protein) gene of P...
2017: Frontiers in Microbiology
Kitikarn Sakuna, Jennifer Elliman, Leigh Owens
Chequa iflavirus (+ve sense ssRNA virus) infects redclaw crayfish (Cherax quadricarinatus) and it may cause mortality reaching 20-40% after about three weeks following stress. The sequence of the RNA-dependent RNA polymerase at nucleotide position 8383-9873 was used for developing and comparing PCR-based detection protocols. The reverse transcription, quantitative, polymerase chain reaction (RT-qPCR) was specific against nine Picornavirales and crustacean viruses and its' measurement of uncertainty (0.07-1...
January 2018: Journal of Virological Methods
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