scholarly journals A novel loop-mediated isothermal amplification-lateral-flow-dipstick (LAMP-LFD) device for rapid detection of Toxoplasma gondii in the blood of stray cats and dogs

Parasite ◽  
2021 ◽  
Vol 28 ◽  
pp. 41
Author(s):  
Yangji Xue ◽  
Qingming Kong ◽  
Haojie Ding ◽  
Chengzuo Xie ◽  
Bin Zheng ◽  
...  
Keyword(s):  
Toxoplasma Gondii ◽  
Lateral Flow ◽  
Isothermal Amplification ◽  
Diagnostic Methods ◽  
Molecular Diagnostic ◽  
Diagnostic Tools ◽  
Loop Mediated Isothermal Amplification ◽  
Stray Cats ◽  
Lateral Flow Dipstick ◽  
Parasitic Pathogens

Toxoplasma gondii is an obligate intracellular protozoan parasite that causes toxoplasmosis and threatens warm-blooded animal and human health worldwide. Simple and applicable diagnostic methods are urgently needed to guide development of effective approaches for prevention of toxoplasmosis. Most molecular diagnostic tools for T. gondii infection require high technical skills, sophisticated equipment, and a controlled lab environment. In this study, we developed a loop-mediated isothermal amplification-lateral-flow-dipstick (LAMP-LFD) assay that specifically targets the 529 bp for detecting T. gondii infection. This novel portable device is universal, fast, user-friendly, and guarantees experimental sensitivity as well as low risk of aerosol contamination. Our LAMP-LFD assay has a detection limit of 1 fg of T. gondii DNA, and shows no cross-reaction with other parasitic pathogens, including Cryptosporidium parvum, Leishmania donovani, and Plasmodium vivax. We validated the developed assay by detecting T. gondii in DNA extracted from blood samples collected from 318 stray cats and dogs sampled from Deqing, Wenzhou, Yiwu, Lishui and Zhoushan cities across Zhejiang province, Eastern China. The LAMP-LFD device detected T. gondii DNA in 4.76 and 4.69% of stray cats and dogs, respectively. In conclusion, the developed LAMP-LFD assay is efficient, minimizes aerosol contamination, and is therefore suitable for detecting T. gondii across basic medical institutions and field settings.

Rapid Detection of Toxoplasma gondii: Prevalence Investigation of T. gondii Infection Among Stray Cats and Dogs in Zhejiang Province Based on a New Loop-Mediated Isothermal Amplification-Lateral-Flow-Dipstick (LAMP-LFD) Device

2020 ◽  
Author(s):  
Yangji Xue ◽  
Shaohong Lu ◽  
Qingming Kong ◽  
Haojie Ding ◽  
Jianzu Ding ◽  
...  
Keyword(s):  
Toxoplasma Gondii ◽  
Lateral Flow ◽  
Zhejiang Province ◽  
Isothermal Amplification ◽  
Diagnostic Methods ◽  
Infection Prevalence ◽  
Loop Mediated Isothermal Amplification ◽  
Stray Cats ◽  
Lateral Flow Dipstick ◽  
Parasitic Pathogens

Abstract Background: Toxoplasma gondii (T. gondii) is worldwide spread caused Toxoplasmosis threatening warm-blooded animal and human health, especially for immunodeficient population and pregnant women. Simple and applicable diagnostic methods are urgently needed for the prevention of toxoplasmosis. The molecular diagnosis of T. gondii infection generally requires high technical skills, sophisticated equipment and a controlled lab environment.Methods: In this study, we developed a loop-mediated isothermal amplification-lateral-flow-dipstick (LAMP-LFD) assay that specifically targets the 529 bp for the detection of T. gondii infection in a new kind of portable device, which is universal, fast, user-friendliness, experimental sensitivity and low risk of aerosol contamination. Results: The detection limit of the LAMP-LFD assay is 1 fg of T. gondii DNA and no cross-reaction with other parasitic pathogens including Leishmania donovani, Plasmodium vivax, Cryptosporidium parvum, etc. In total, 318 stray cat and dog blood samples were collected from Deqing, Wenzhou, Yiwu, Lishui and Zhoushan cities in Zhejiang province, Eastern China. The current infection prevalence of T. gondii was 4.76% and 4.69% in stray cats and dogs respectively, detected by LAMP-LFD device.Conclusions: In conclusion, the established LAMP-LFD was an efficient and avoidable aerosol-contaminated device that can detect 1 fg genomic DNA of T. gondii, and suitable for T. gondii detection in the basic medical institution and even in field areas.

scholarly journals Lateral Flow Loop-Mediated Isothermal Amplification Test with Stem Primers: Detection ofCryptosporidiumSpecies in Kenyan Children Presenting with Diarrhea

2018 ◽  
Vol 2018 ◽  
pp. 1-9 ◽  
Author(s):  
Timothy S. Mamba ◽  
Cecilia K. Mbae ◽  
Johnson Kinyua ◽  
Erastus Mulinge ◽  
Gitonga Nkanata Mburugu ◽  
...  
Keyword(s):  
Nested Pcr ◽  
Lateral Flow ◽  
Isothermal Amplification ◽  
Diagnostic Methods ◽  
Diagnostic Tools ◽  
Clinical Samples ◽  
Analytical Sensitivity ◽  
Flow Loop ◽  
Loop Mediated Isothermal Amplification ◽  
Higher Sensitivity

Background. Cryptosporidiumis a protozoan parasite and a major cause of diarrhea in children and immunocompromised patients. Current diagnostic methods for cryptosporidiosis such as microscopy have low sensitivity while techniques such as PCR indicate higher sensitivity levels but are seldom used in developing countries due to their associated cost. A loop-mediated isothermal amplification (LAMP) technique, a method with shorter time to result and with equal or higher sensitivity compared to PCR, has been developed and applied in the detection ofCryptosporidiumspecies. The test has a detection limit of 10 pg/µl (~100 oocysts/ml) indicating a need for more sensitive diagnostic tools. This study developed a more sensitive lateral flow dipstick (LFD) LAMP test based on SAM-1 gene and with the addition of a second set of reaction accelerating primers (stem primers).Results. The stem LFD LAMP test showed analytical sensitivity of 10 oocysts/ml compared to 100 oocysts/ml (10 pg/ul) for each of the SAM-1 LAMP test and nested PCR. The stem LFD LAMP and nested PCR detected 29/39 and 25/39 positive samples of previously identifiedC. parvumandC. hominisDNA, respectively. The SAM-1 LAMP detected 27/39. On detection ofCryptosporidiumDNA in 67 clinical samples, the stem LFD LAMP detected 16 samples and SAM-2 LAMP 14 and nested PCR identified 11. Preheating the templates increased detection by stem LFD LAMP to 19 samples. Time to results from master mix preparation step took ~80 minutes. The test was specific, and no cross-amplification was recorded with nontarget DNA.Conclusion.The developed stem LFD LAMP test is an appropriate method for the detection ofC. hominis, C. parvum,andC. meleagridisDNA in human stool samples. It can be used in algorithm with other diagnostic tests and may offer promise as an effective diagnostic tool in the control of cryptosporidiosis.

scholarly journals Development of a loop-mediated isothermal amplification (LAMP) assay for the identification of the invasive wood borer Aromia bungii (Coleoptera: Cerambycidae) from frass

3 Biotech ◽  
2021 ◽  
Vol 11 (2) ◽  
Author(s):  
Domenico Rizzo ◽  
Nicola Luchi ◽  
Daniele Da Lio ◽  
Linda Bartolini ◽  
Francesco Nugnes ◽  
...  
Keyword(s):  
Lamp Assay ◽  
Isothermal Amplification ◽  
Molecular Diagnostic ◽  
Diagnostic Tools ◽  
Loop Mediated Isothermal Amplification ◽  
Larval Stages ◽  
Longhorn Beetle ◽  
Rapid Monitoring ◽  
Major Pest ◽  
Wood Borer

AbstractThe red-necked longhorn beetle Aromia bungii (Faldermann, 1835) (Coleoptera: Cerambycidae) is native to east Asia, where it is a major pest of cultivated and ornamental species of the genus Prunus. Morphological or molecular discrimination of adults or larval specimens is required to identify this invasive wood borer. However, recovering larval stages of the pest from trunks and branches causes extensive damage to plants and is timewasting. An alternative approach consists in applying non-invasive molecular diagnostic tools to biological traces (i.e., fecal pellets, frass). In this way, infestations in host plants can be detected without destructive methods. This paper presents a protocol based on both real-time and visual loop-mediated isothermal amplification (LAMP), using DNA of A. bungii extracted from fecal particles in larval frass. Laboratory validations demonstrated the robustness of the protocols adopted and their reliability was confirmed performing an inter-lab blind panel. The LAMP assay and the qPCR SYBR Green method using the F3/B3 LAMP external primers were equally sensitive, and both were more sensitive than the conventional PCR (sensitivity > 103 to the same starting matrix). The visual LAMP protocol, due to the relatively easy performance of the method, could be a useful tool to apply in rapid monitoring of A. bungii and in the management of its outbreaks.

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.

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