scholarly journals Simple, sensitive and cost-effective detection of wAlbB Wolbachia in Aedes mosquitoes, using loop mediated isothermal amplification combined with the electrochemical biosensing method

2021 ◽  
Author(s):  
Pattamaporn Kittayapong ◽  
Parinda Thayanukul ◽  
Benchaporn Lertanantawong ◽  
Worachart Sirawaraporn ◽  
Surat Charasmongkolcharoen ◽  
...  
Keyword(s):  
Low Cost ◽  
High Sensitivity ◽  
Cost Effective ◽  
Isothermal Amplification ◽  
Loop Mediated Isothermal Amplification ◽  
Control Programs ◽  
Electrochemical Biosensing ◽  
Dna Strand Displacement ◽  
Displacement Based ◽  
Dna Strand

Background Wolbachia is an endosymbiont bacterium generally found in about 40% of insects, including mosquitoes, but it is absent in Aedes aegypti which is an important vector of several arboviral diseasesdengue, chikungunya, Zika, and yellow fever. The evidence that Wolbachia trans-infected Ae. aegypti mosquitoes lost their vectorial competence and became less capable of transmitting arboviruses to human hosts highlights the potential of using Wolbachia - based approaches for prevention and control of arboviral diseases. Recently, release of Wolbachia trans-infected Ae. aegypti has been deployed widely in many countries for the control of mosquito-borne viral diseases. Field surveillance and monitoring of Wolbachia presence in released mosquitoes is important for the success of these control programs. So far, a number of studies have reported the development of loop mediated isothermal amplification (LAMP) assays to detect Wolbachia in mosquitoes, but the methods still have some specificity issues. Methodology/Principal Findings We describe here the development of a LAMP combined with the DNA strand displacement-based electrochemical sensor (BIOSENSOR) method to detect wAlbB Wolbachia in trans-infected Ae . aegypti . Our developed LAMP primers were more specific to wAlbB detection than those of the previous published ones if  the assays were conducted with low-cost and non-specific detecting dyes. The detection capacity of our LAMP technique was 3.8 nM and the detection limit reduced to 2.16 fM when combined with the BIOSENSOR. Our study demonstrates that the BIOSENSOR can also be applied as a stand-alone method for detecting Wolbachia ; and it showed high sensitivity when used with the crude DNA extracts of macerated mosquito samples without DNA purification. Conclusions/Significance Our results suggest that both LAMP and BIOSENSOR, either used in combination or stand-alone, are robust and sensitive. The methods have good potential for routine detection of Wolbachia in mosquitoes during field surveillance and monitoring of Wolbachia -based release programs, especially in countries with limited resources.

scholarly journals A Rapid, Sensitive, Low-Cost Assay for Detecting Hydrogenotrophic Methanogens in Anaerobic Digesters Using Loop-Mediated Isothermal Amplification

2020 ◽  
Vol 8 (5) ◽  
pp. 740
Author(s):  
Anna M. Alessi ◽  
Bing Tao ◽  
Wei Zhang ◽  
Yue Zhang ◽  
Sonia Heaven ◽  
...  
Keyword(s):  
Low Cost ◽  
Quantitative Measure ◽  
Methanogenic Archaea ◽  
High Sensitivity ◽  
Gas Production ◽  
Isothermal Amplification ◽  
Process Conditions ◽  
Strictly Anaerobic ◽  
Loop Mediated Isothermal Amplification ◽  
Hydrogenotrophic Methanogens

Understanding how the presence, absence, and abundance of different microbial genera supply specific metabolic functions for anaerobic digestion (AD) and how these impact on gas production is critical for a long-term understanding and optimization of the AD process. The strictly anaerobic methanogenic archaea are essential for methane production within AD microbial communities. Methanogens are a phylogenetically diverse group that can be classified into three metabolically distinct lineages based on the substrates they use to produce methane. While process optimization based on physicochemical parameters is well established in AD, measurements that could allow manipulation of the underlying microbial community are seldom used as they tend to be non-specific, expensive, or time-consuming, or a combination of all three. Loop-mediated isothermal amplification (LAMP) assays combine a simple, rapid, low-cost detection technique with high sensitivity and specificity. Here, we describe the optimization of LAMP assays for the detection of four different genera of hydrogenotrophic methanogens: Methanoculleus, Methanothermobacter, Methanococcus, and Methanobrevibacter spp. By targeting archaeal elongation factor 2 (aEF2), these LAMP assays provide a rapid, low-cost, presence/absence indication of hydrogenotrophic methanogens that could be used as a real-time measure of process conditions. The assays were shown to be sensitive to 1 pg of DNA from most tested methanogen species, providing a route to a quantitative measure through simple serial dilution of samples. The LAMP assays described here offer a simple, fast, and affordable method for the specific detection of four different genera of hydrogenotrophic methanogens. Our results indicate that this approach could be developed into a quantitative measure that could provide rapid, low-cost insight into the functioning and optimization of AD and related systems.

scholarly journals Development of Detection Equipment for a Polymerase Chain Reaction with a Loop-Mediated Isothermal Amplification Reaction

2021 ◽  
Vol 2021 ◽  
pp. 1-9
Author(s):  
Wei-Chien Weng ◽  
Yu-Cheng Lin
Keyword(s):  
Polymerase Chain Reaction ◽  
Temperature Control ◽  
Low Cost ◽  
Cost Effective ◽  
Isothermal Amplification ◽  
Raspberry Pi ◽  
Chain Reaction ◽  
Loop Mediated Isothermal Amplification ◽  
Processing Signal ◽  
Polymerase Chain

In this research, low-cost detection equipment intended to carry out a polymerase chain reaction (PCR) through a loop-mediated isothermal amplification (LAMP) reaction is presented. We designed the internal structure with SolidWorks and AutoCAD. The equipment comprised a Raspberry Pi development board, a temperature control module, and a fluorescent optical detection module. The main program, temperature control, florescent signal processing, signal analysis, and screen display were programmed with Java. We applied the digital temperature controller module to obtain precise temperature control of the equipment. The experimental results showed that the heating rate of the testing equipment could reach 65°C within 4 minutes and could be accurately controlled to within 1°C. The duration of the LAMP PCR experiment was found to be significantly shorter than that of the conventional PCR. The results also revealed that with LAMP PCR, the temperature could be accurately controlled within a specific range, and the designed heating tasks could be completed within 15 minutes to one hour, depending on the specimen. The equipment could also correctly read both the positive and negative reactions with fluorescent signals. Thus, the proposed LAMP PCR detection equipment is more sensitive, more stable, and more cost-effective than other conventional alternatives and can be used in numerous clinical applications.

scholarly journals SARS-CoV-2 Direct Detection Without RNA Isolation With Loop-Mediated Isothermal Amplification (LAMP) and CRISPR-Cas12

2021 ◽  
Vol 8 ◽  
Author(s):  
Alfredo Garcia-Venzor ◽  
Bertha Rueda-Zarazua ◽  
Eduardo Marquez-Garcia ◽  
Vilma Maldonado ◽  
Angelica Moncada-Morales ◽  
...  
Keyword(s):  
Limit Of Detection ◽  
Direct Detection ◽  
Direct Method ◽  
Rna Extraction ◽  
Rna Isolation ◽  
High Sensitivity ◽  
High Specificity ◽  
Isothermal Amplification ◽  
Clinical Samples ◽  
Loop Mediated Isothermal Amplification

As to date, more than 49 million confirmed cases of Coronavirus Disease 19 (COVID-19) have been reported worldwide. Current diagnostic protocols use qRT-PCR for viral RNA detection, which is expensive and requires sophisticated equipment, trained personnel and previous RNA extraction. For this reason, we need a faster, direct and more versatile detection method for better epidemiological management of the COVID-19 outbreak. In this work, we propose a direct method without RNA extraction, based on the Loop-mediated isothermal amplification (LAMP) and Clustered Regularly Interspaced Short Palindromic Repeats-CRISPR associated protein (CRISPR-Cas12) technique that allows the fast detection of SARS-CoV-2 from patient samples with high sensitivity and specificity. We obtained a limit of detection of 16 copies/μL with high specificity and at an affordable cost. The diagnostic test readout can be done with a real-time PCR thermocycler or with the naked eye in a blue-light transilluminator. Our method has been evaluated on a small set of clinical samples with promising results.

scholarly journals Loop-Mediated Isothermal Amplification for Detection ofStaphylococcus aureusin Dairy Cow Suffering from Mastitis

2012 ◽  
Vol 2012 ◽  
pp. 1-5 ◽  
Author(s):  
Zhang Tie ◽  
Wang Chunguang ◽  
Wei Xiaoyuan ◽  
Zhao Xinghua ◽  
Zhong Xiuhui
Keyword(s):  
Staphylococcus Aureus ◽  
High Sensitivity ◽  
High Specificity ◽  
Isothermal Amplification ◽  
Lamp Method ◽  
Specific Primers ◽  
Loop Mediated Isothermal Amplification ◽  
Reaction Tube ◽  
Specificity And Sensitivity ◽  
Detectable Concentration

To develop a rapid detection method ofStaphylococcus aureususing loop-mediated isothermal amplification (LAMP), four specific primers were designed according to six distinct sequences of thenucgene. In addition, the specificity and sensitivity of LAMP were verified and compared with those of PCR. Results showed that the LAMP reaction was completed within 45 min at 62.5°C, and ladder bands were appeared in LAMP products analyzed by gel electrophoresis. After adding 1x SYBR Green l, the positive reaction tube showed green color and the negative reaction tube remained orange, indicating that the LAMP has high specificity. The minimal detectable concentration of LAMP was1×102 CFU/mL and that of PCR was1×104 CFU/mL, indicating that the LAMP was 100 times more sensitive than the PCR. The LAMP method for detection ofStaphylococcus aureushas many advantages, such as simple operation, high sensitivity, high specificity, and rapid analysis. Therefore, this method is more suitable for the rapid on-site detection ofStaphylococcus aureus.

scholarly journals Loop-mediated isothermal amplification (LAMP) : A rapid molecular diagnosis technique for detection of human pathogens

2017 ◽  
Vol 07 (03) ◽  
pp. 042-048
Author(s):  
Gunimala Chakraborty ◽  
Indrani Karunasagar ◽  
Anirban Chakraborty
Keyword(s):  
Treatment Strategies ◽  
Lamp Assay ◽  
Cost Effective ◽  
Isothermal Amplification ◽  
Current Status ◽  
Molecular Diagnostic ◽  
Diagnostic Tools ◽  
Human Pathogens ◽  
Quality Healthcare ◽  
Loop Mediated Isothermal Amplification

AbstractDelivery of quality healthcare in case of an infectious disease depends on how efficiently and how quickly the responsible pathogens are detected from the samples. Molecular methods can detect the presence of pathogens in a rapid and sensitive manner. Over the years, a number of such assays have been developed. However, these methods, although highly reliable and efficient, require use of expensive equipment, reagents, and trained personnel. Therefore, development of molecular assays that are simple, rapid, cost-effective, yet sensitive, is highly warranted to ensure efficient management or treatment strategies. Loop-mediated isothermal amplification (LAMP), a technique invented in the year 2000, is a novel method that amplifies DNA at isothermal conditions. Since its invention, this technique has been one of the most extensively used molecular diagnostic tools in the field of diagnostics offering rapid, accurate and cost-effective diagnosis of infectious diseases. Using the LAMP principle, many commercial kits have been developed in the last decade for a variety of human pathogens including bacteria, viruses and parasites. Currently LAMP assay is being considered as an effective diagnostic tool for use in developing countries because of its simple working protocol, allowing even an onsite application. The focus of this review is to describe the salient features of this technique the current status of development of LAMP assays with an emphasis on the pathogens of clinical significance.

Colorimetric loop-mediated isothermal amplification (LAMP) for cost-effective and quantitative detection of SARS-CoV-2: the change in color in LAMP-based assays quantitatively correlates with viral copy number

2021 ◽  
Author(s):  
Everardo González-González ◽  
Itzel Montserrat Lara-Mayorga ◽  
Iram Pablo Rodríguez-Sánchez ◽  
Yu Shrike Zhang ◽  
Sergio O. Martínez-Chapa ◽  
...  
Keyword(s):  
Copy Number ◽  
Quantitative Method ◽  
Diagnostic Performance ◽  
Cost Effective ◽  
Isothermal Amplification ◽  
Quantitative Detection ◽  
Loop Mediated Isothermal Amplification ◽  
Viral Copy Number ◽  
Viral Copy

Colorimetric LAMP for COVID-19 intensified diagnostics: a simple and quantitative method comparable in diagnostic performance to RT-qPCR.

scholarly journals Artificial Intelligence-Assisted Loop Mediated Isothermal Amplification (AI-LAMP) for Rapid Detection of SARS-CoV-2

Viruses ◽  
2020 ◽  
Vol 12 (9) ◽  
pp. 972 ◽  
Author(s):  
Mohammed A. Rohaim ◽  
Emily Clayton ◽  
Irem Sahin ◽  
Julianne Vilela ◽  
Manar E. Khalifa ◽  
...  
Keyword(s):  
Artificial Intelligence ◽  
De Novo ◽  
Tracking System ◽  
Lamp Assay ◽  
Cost Effective ◽  
Isothermal Amplification ◽  
Analytical Sensitivity ◽  
Diagnostic Device ◽  
Loop Mediated Isothermal Amplification ◽  
Resource Limited

Until vaccines and effective therapeutics become available, the practical solution to transit safely out of the current coronavirus disease 19 (CoVID-19) lockdown may include the implementation of an effective testing, tracing and tracking system. However, this requires a reliable and clinically validated diagnostic platform for the sensitive and specific identification of SARS-CoV-2. Here, we report on the development of a de novo, high-resolution and comparative genomics guided reverse-transcribed loop-mediated isothermal amplification (LAMP) assay. To further enhance the assay performance and to remove any subjectivity associated with operator interpretation of results, we engineered a novel hand-held smart diagnostic device. The robust diagnostic device was further furnished with automated image acquisition and processing algorithms and the collated data was processed through artificial intelligence (AI) pipelines to further reduce the assay run time and the subjectivity of the colorimetric LAMP detection. This advanced AI algorithm-implemented LAMP (ai-LAMP) assay, targeting the RNA-dependent RNA polymerase gene, showed high analytical sensitivity and specificity for SARS-CoV-2. A total of ~200 coronavirus disease (CoVID-19)-suspected NHS patient samples were tested using the platform and it was shown to be reliable, highly specific and significantly more sensitive than the current gold standard qRT-PCR. Therefore, this system could provide an efficient and cost-effective platform to detect SARS-CoV-2 in resource-limited laboratories.

scholarly journals Development of an in-house loop-mediated isothermal amplification (LAMP) assay for detection of Mycobacterium tuberculosis and evaluation in sputum samples of Nepalese patients

2008 ◽  
Vol 57 (4) ◽  
pp. 439-443 ◽  
Author(s):  
Basu Dev Pandey ◽  
Ajay Poudel ◽  
Tomoko Yoda ◽  
Aki Tamaru ◽  
Naozumi Oda ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Lamp Assay ◽  
High Sensitivity ◽  
Isothermal Amplification ◽  
Nucleic Acid Amplification ◽  
Lamp Method ◽  
Rrna Gene ◽  
Predictive Values ◽  
Loop Mediated Isothermal Amplification ◽  
System A

A number of nucleic acid amplification assays (NAAs) have been employed to detect tubercle bacilli in clinical specimens for tuberculosis (TB) diagnosis. Among these, loop-mediated isothermal amplification (LAMP) is an NAA possessing superior isothermal reaction characteristics. In the present study, a set of six specific primers targeting the Mycobacterium tuberculosis 16S rRNA gene with high sensitivity was selected and a LAMP system (MTB-LAMP) was developed. Using this system, a total of 200 sputum samples from Nepalese patients were investigated. The sensitivity of MTB-LAMP in culture-positive samples was 100 % (96/96), and the specificity in culture-negative samples was 94.2 % (98/104, 95 % confidence interval 90.5–97.9 %). The positive and negative predictive values of MTB-LAMP were 94.1 and 100 %, respectively. These results indicate that this MTB-LAMP method may prove to be a powerful tool for the early diagnosis of TB.

scholarly journals Loop Mediated Isothermal Amplification: Principles and Applications in Plant Virology

Plants ◽  
2020 ◽  
Vol 9 (4) ◽  
pp. 461 ◽  
Author(s):  
Stefano Panno ◽  
Slavica Matić ◽  
Antonio Tiberini ◽  
Andrea Giovanni Caruso ◽  
Patrizia Bella ◽  
...  
Keyword(s):  
Plant Protection ◽  
Point Of Care ◽  
Low Cost ◽  
Isothermal Amplification ◽  
Nucleic Acid Amplification ◽  
Plant Virology ◽  
Food Industries ◽  
Loop Mediated Isothermal Amplification ◽  
Application Fields ◽  
Agriculture And Food

In the last decades, the evolution of molecular diagnosis methods has generated different advanced tools, like loop-mediated isothermal amplification (LAMP). Currently, it is a well-established technique, applied in different fields, such as the medicine, agriculture, and food industries, owing to its simplicity, specificity, rapidity, and low-cost efforts. LAMP is a nucleic acid amplification under isothermal conditions, which is highly compatible with point-of-care (POC) analysis and has the potential to improve the diagnosis in plant protection. The great advantages of LAMP have led to several upgrades in order to implement the technique. In this review, the authors provide an overview reporting in detail the different LAMP steps, focusing on designing and main characteristics of the primer set, different methods of result visualization, evolution and different application fields, reporting in detail LAMP application in plant virology, and the main advantages of the use of this technique.

scholarly journals Loop-Mediated Isothermal Amplification as Point-of-Care Diagnosis for Neglected Parasitic Infections

2020 ◽  
Vol 21 (21) ◽  
pp. 7981
Author(s):  
Catalina Avendaño ◽  
Manuel Alfonso Patarroyo
Keyword(s):  
Neglected Tropical Diseases ◽  
Point Of Care ◽  
Low Cost ◽  
High Specificity ◽  
World Health Organisation ◽  
Isothermal Amplification ◽  
World Health ◽  
Parasitic Infections ◽  
River Blindness ◽  
Loop Mediated Isothermal Amplification

The World Health Organisation (WHO) has placed twenty diseases into a group known as neglected tropical diseases (NTDs), twelve of them being parasitic diseases: Chagas’ disease, cysticercosis/taeniasis, echinococcosis, food-borne trematodiasis, human African trypanosomiasis (sleeping sickness), leishmaniasis, lymphatic filariasis, onchocerciasis (river blindness), schistosomiasis, soil-transmitted helminthiasis (ascariasis, hookworm, trichuriasis), guinea-worm and scabies. Such diseases affect millions of people in developing countries where one of the main problems concerning the control of these diseases is diagnosis-based due to the most affected areas usually being far from laboratories having suitable infrastructure and/or being equipped with sophisticated equipment. Advances have been made during the last two decades regarding standardising and introducing techniques enabling diagnoses to be made in remote places, i.e., the loop-mediated isothermal amplification (LAMP) technique. This technique’s advantages include being able to perform it using simple equipment, diagnosis made directly in the field, low cost of each test and the technique’s high specificity. Using this technique could thus contribute toward neglected parasite infection (NPI) control and eradication programmes. This review describes the advances made to date regarding LAMP tests, as it has been found that even though several studies have been conducted concerning most NPI, information is scarce for others.

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