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loop-mediated isothermal amplification

Yohei Kurosaki, Sayaka Okada, Sayuri Nakamae, Jiro Yasuda
Bovine papular stomatitis virus (BPSV) causes pustular cutaneous disease in cattle worldwide. This paper describes the development of a specific loop-mediated isothermal amplification (LAMP) assay to detect BPSV which did not cross-react with other parapoxviruses. To assess analytical sensitivity of this LAMP assay, DNA was extracted from serially diluted BPSV from which the infectious titer was determined by a novel assay based on calf kidney epithelial cells. The LAMP assay had equivalent analytical sensitivity to quantitative PCR, and could detect as few as 86 copies of viral DNA per reaction...
October 14, 2016: Journal of Virological Methods
Islam I Sabike, Ryoko Uemura, Yumi Kirino, Hirohisa Mekata, Satoshi Sekiguchi, Tamaki Okabayashi, Yoshitaka Goto, Wataru Yamazaki
Rapid identification of Campylobacter-positive flocks before slaughter, following freezing and heat treatment for the Campylobacter-positive carcasses at the slaughterhouses is an effective control strategy against foodborne campylobacteriosis. We evaluated a loop-mediated isothermal amplification (LAMP) assay for the direct screening of naturally contaminated chicken cloacal swabs for C. jejuni/C. coli to compare this assay with conventional quantitative culture methods. In a comparison study of 165 broilers, the LAMP assay showed 82...
2016: Frontiers in Microbiology
Mohammad Amin Almasi, Galavizh Almasi
Sugar beet can be infected by many different viruses that can reduce yield; beet necrotic yellow vein virus (BNYVV) is one of the most economically important viruses of this crop plant. This report describes a new reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay for identification of BNYVV. In addition, a novel immunocapture (IC) RT-LAMP assay for rapid and easy detection (without RNA extraction) of BNYVV was developed here and compared with DAS-ELISA and RT-LAMP assays. Our results show that the IC-RT-LAMP assay is a highly reliable alternative assay for identification of BNYVV...
October 13, 2016: Archives of Virology
Xiangfeng He, Fei Xue, Shufa Xu, Wenhe Wang
Lily symptomless virus (LSV) is one of the most prevalent viruses that infect lily plants worldwide. A rapid and sensitive reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay was developed for detection of LSV, using two primer pairs that specifically amplified the conserved sequence of LSV coat protein. The optimum reaction conditions were as follows: 4mM MgCl2 and 0.8M betaine with incubation at 64°C for 30min. The limit of detection of LSV from infected lily leaves was 10-fold higher for RT-LAMP than for conventional RT-PCR...
October 10, 2016: Journal of Virological Methods
Yuka Okuwa, Michiko Miyamato-Hayashi, Takaichi Tanaka, Yuji Hayakawa, Yasuo Inoshima
Various techniques for screening and detection of bovine leukemia virus (BLV) were compared to ascertain a rapid and simple technique for routine examination. The performance of real-time PCR, nested PCR and loop-mediated isothermal amplification (LAMP) assays was compared using DNA extracted from whole blood instead of white blood cells (WBCs) of 23 cattle. Real-time PCR, LAMP and nested PCR detected 18, 16 and 11 BLV-positive cattle, respectively. These results suggest that LAMP using DNA from whole blood could enable rapid examination, as isolation of WBCs and electrophoresis is time-consuming and could be useful as a simple, and rapid method for routine screening of BLV...
October 3, 2016: Journal of Veterinary Medical Science
Gaolian Xu, Hang Zhao, Jonathan M Cooper, Julien Reboud
We demonstrate a multiplexed loop mediated isothermal amplification (LAMP) assay for infectious disease diagnostics, where the analytical process flow of target pathogens genomic DNA is performed manually by moving magnetic beads through a series of plugs in a capillary. Heat is provided by a water bath and the results are read by the naked eye, enabling applications in low resource settings.
October 6, 2016: Chemical Communications: Chem Comm
Gihoon Choi, Daniel Song, Sony Shrestha, Jun Miao, Liwang Cui, Weihua Guan
In response to the urgent need of a field-deployable and highly sensitive malaria diagnosis, we developed a standalone, "sample-in-answer-out" molecular diagnostic system (AnyMDx) to enable quantitative molecular analysis of blood-borne malaria in low resource areas. The system consists of a durable battery-powered analyzer and a disposable microfluidic compact disc loaded with reagents ready for use. A low power thermal module and a novel fluorescence-sensing module are integrated into the analyzer for real-time monitoring of loop-mediated isothermal nucleic acid amplification (LAMP) of target parasite DNA...
October 10, 2016: Lab on a Chip
Wenfang Du, Mengmei Lv, Junjie Li, Ruqin Yu, Jianhui Jiang
A novel ligation-based loop-mediated isothermal amplification (ligation-LAMP) method has been developed for miRNA detection, which enables highly selective and sensitive quantitative detection of miR-21 in a dynamic range from 1 fM to 1 nM with the ability to discriminate single-base mismatches.
October 10, 2016: Chemical Communications: Chem Comm
Tian-Min Qiao, Jing Zhang, Shu-Jiang Li, Shan Han, Tian-Hui Zhu
Eucalyptus dieback disease, caused by Cylindrocladium scoparium, has occurred in last few years in large Eucalyptus planting areas in China and other countries. Rapid, simple, and reliable diagnostic techniques are desired for the early detection of Eucalyptus dieback of C. scoparium prior to formulation of efficient control plan. For this purpose, three PCR-based methods of nested PCR, multiplex PCR, loop-mediated isothermal amplification (LAMP) were developed for detection of C. scoparium based on factor 1-alpha (tef1) and beta-tubulin gene in this study...
October 2016: Plant Pathology Journal
Robert D Stedtfeld, Tiffany M Stedtfeld, Farag Samhan, Yogendra H Kanitkar, Paul B Hatzinger, Alison M Cupples, Syed A Hashsham
Nucleic acid amplification of biomarkers is increasingly used to monitor microbial activity and assess remedial performance in contaminated aquifers. Previous studies described the use of filtration, elution, and direct isothermal amplification (i.e. no DNA extraction and purification) as a field-able means to quantify Dehalococcoides spp. in groundwater. This study expands previous work with direct loop mediated isothermal amplification (LAMP) for the detection and quantification of Dehalobacter spp. in groundwater...
October 5, 2016: Journal of Microbiological Methods
Mary Prahl, Prasanna Jagannathan, Tara I McIntyre, Ann Auma, Lila Farrington, Samuel Wamala, Mayimuna Nalubega, Kenneth Musinguzi, Kate Naluwu, Esther Sikyoma, Rachel Budker, Hilary Vance, Pamela Odorizzi, Patience Nayebare, John Ategeka, Abel Kakuru, Diane V Havlir, Moses R Kamya, Grant Dorsey, Margaret E Feeney
BACKGROUND: In malaria-endemic areas, the first exposure to malaria antigens often occurs in utero when the fetal immune system is poised towards the development of tolerance. Children exposed to placental malaria have an increased risk of clinical malaria in the first few years of life compared to unexposed children. Recent work has suggested the potential of pregnancy-associated malaria to induce immune tolerance in children living in malaria-endemic areas. A study was completed to evaluate the effect of malaria exposure during pregnancy on fetal immune tolerance and effector responses...
October 7, 2016: Malaria Journal
Estefanía J Valverde, Irene Cano, Dolores Castro, Richard K Paley, Juan J Borrego
Lymphocystis disease virus (LCDV) infections have been described in gilthead seabream (Sparus aurata L.) and Senegalese sole (Solea senegalensis, Kaup), two of the most important marine fish species in the Mediterranean aquaculture. In this study, a rapid, specific, and sensitive detection method for LCDV genotype VII based on loop-mediated isothermal amplification (LAMP) was developed. The LAMP assay, performed using an apparatus with real-time amplification monitoring, was able to specifically detect LCDV genotype VII from clinically positive samples in less than 12 min...
October 5, 2016: Food and Environmental Virology
Zeynep Koloren, Onuralp Seferoğlu, Panagiotis Karanis
A total of 420 environmental water samples and 120 drinking water samples from 45 different sampling sites of the Black Sea in Turkey were collected between 2012 and 2014. Genomic DNA was isolated from all the investigated water samples and comparativelly analyzed by Loop-mediated isothermal amplification (LAMP) of the elongation factor 1 Alfa (EF1α) gene, and by nested Polymerase Chain Reaction (nPCR) of the small subunit (SSU) rRNA and semi-nested PCR (snPCR) of the glutamate dehydrogenase gene (GDH). 141 (58...
September 30, 2016: Acta Tropica
Ilada Choopara, Narong Arunrut, Wansika Kiatpathomchai, Deborah Dean, Naraporn Somboonna
We developed an assay comprising crude DNA lysis by simple heat treatment coupled loop-mediated isothermal amplification with hydroxynaphthol blue (LAMP-HNB) for C. trachomatis detection, and evaluated the developed assay for its feasibility as one-step point-of-care detection on 284 endocervical swab specimens from clinically symptomatic C. trachomatis and healthy subjects. This assay is sensitive to 0.04 pg of ompA, specific with 6 primers targeting C. trachomatis ompA region, rapid (45 min total assay time), inexpensive (~3 USD/reaction), does not require sophisticated instrumentation, and has comparable assay effectiveness (95% specificity, 90-100% sensitivity) to the bacterial DNA isolation by commercial kit coupled with PCR and gel electrophoresis (98-100% specificity, 87-100% sensitivity) based on the clinical samples test...
September 30, 2016: Letters in Applied Microbiology
El Mehdi Bentaleb, Mohammed Abid, My Driss El Messaoudi, Brahim Lakssir, El Mostafa Ressami, Saaïd Amzazi, Hassan Sefrioui, Hassan Ait Benhassou
BACKGROUND: Tuberculosis (TB) is a major global health problem and remains the leading cause of morbidity and mortality in developing countries. Routinely used TB diagnostic methods, in most endemic areas, are time-consuming, often less-sensitive, expensive and inaccessible to most patients. Therefore, there is an urgent need for the development of early, easy to use and effective diagnosis tools of TB, which can be effectively integrated into resource limited settings, to anticipate the early treatment and limit further spread of the disease...
September 27, 2016: BMC Infectious Diseases
Damien M O'Halloran
Overlapping PCR is commonly used in many molecular applications that include stitching PCR fragments together, generating fluorescent transcriptional and translational fusions, inserting mutations, making deletions, and PCR cloning. Overlapping PCR is also used for genotyping and in detection experiments using techniques such as loop-mediated isothermal amplification (LAMP). STITCHER is a web tool providing a central resource for researchers conducting all types of overlapping assembly PCR experiments with an intuitive interface for automated primer design that's fast, easy to use, and freely available online...
2017: Methods in Molecular Biology
Chi-Chu Tsai, Huei-Chuan Shih, Ya-Zhu Ko, Ren-Huang Wang, Shu-Ju Li, Yu-Chung Chiang
Papaya (Carica papaya L.) is an economically important tropical fruit tree with hermaphrodite, male and female sex types. Hermaphroditic plants are the major type used for papaya production because their fruits have more commercial advantages than those of female plants. Sex determination of the seedlings, or during the early growth stages, is very important for the papaya seedling industry. Thus far, the only method for determining the sex type of a papaya at the seedling stage has been DNA analysis. In this study, a molecular technique-based on DNA analysis-was developed for detecting male-hermaphrodite-specific markers to examine the papaya's sex type...
2016: International Journal of Molecular Sciences
S Y Youn, O M Jeong, B K Choi, S C Jung, M S Kang
Raw chicken products are major causes of human foodborne salmonellosis worldwide. In particular, there is a significant risk of human exposure to Salmonella originating from the chicken slaughtering process. Controlling the contamination of chicken carcasses by Salmonella has been a considerable challenge in chicken-slaughtering facilities and involves routine microbiological monitoring using reliable detection methods. Simple and rapid detection methods, particularly those capable of determining cell viability, will significantly facilitate routine monitoring of Salmonella Here, we report an invA-based loop-mediated isothermal amplification method coupled with a simple propidium monoazide treatment (PMA-LAMP) for simple and rapid detection and quantification of viable Salmonella in rinse water of chicken carcasses...
September 24, 2016: Poultry Science
J F C Loo, H C Kwok, C C H Leung, S Y Wu, I L G Law, Y K Cheung, Y Y Cheung, M L Chin, P Kwan, M Hui, S K Kong, H P Ho
Sepsis by bacterial infection causes high mortality in patients in intensive care unit (ICU). Rapid identification of bacterial infection is essential to ensure early appropriate administration of antibiotics to save lives of patients, yet the present benchtop molecular diagnosis is time-consuming and labor-intensive, which limits the treatment efficiency especially when the number of samples to be tested is extensive. Therefore, we hereby report a microfluidic platform lab-on-a-disc (LOAD) to provide a sample-to-answer solution...
September 2, 2016: Biosensors & Bioelectronics
Karin Johansson, Hanna Karlsson, Torbjörn Norén
Diagnostic testing for Clostridium difficile infection (CDI) has, in recent years, seen the introduction of rapid dual-EIA (enzyme immunoassay) tests combining species-specific glutamate dehydrogenase (GDH) with toxin A/B. In a prospective study, we compared the C. DIFF Quik Chek Complete test to a combination of selective culture (SC) and loop-mediated isothermal amplification (LAMP) of the toxin A gene. Of 419 specimens, 68 were positive in SC including 62 positive in LAMP (14.7%). The combined EIA yielded 82 GDH positives of which 47 were confirmed toxin A/B positive (11%) corresponding to a sensitivity and specificity of 94% for GDH EIA compared to SC and for toxin A/B EIA a sensitivity of 71% and a specificity of 99% compared to LAMP...
November 2016: APMIS: Acta Pathologica, Microbiologica, et Immunologica Scandinavica
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