+ Site Statistics
+ Search Articles
+ PDF Full Text Service
How our service works
Request PDF Full Text
+ Follow Us
Follow on Facebook
Follow on Twitter
Follow on LinkedIn
+ Subscribe to Site Feeds
Most Shared
PDF Full Text
+ Translate
+ Recently Requested

Optimization of a Loop Mediated Isothermal Amplification (LAMP) Assay for In-Field Detection of Dichelobacter nodosus With aprV2 (VDN LAMP) in Victorian Sheep Flocks



Optimization of a Loop Mediated Isothermal Amplification (LAMP) Assay for In-Field Detection of Dichelobacter nodosus With aprV2 (VDN LAMP) in Victorian Sheep Flocks



Frontiers in Veterinary Science 6: 67



Dichelobacter nodosus is the primary etiological agent of footrot in sheep and has a variety of virulence factors. Of these, AprV2, an extracellular protease, has been shown to be capable of causing severe or "virulent" disease symptoms under the right conditions. Due to this, a loop-mediated isothermal amplification (LAMP) assay for the detection of aprV2-positive D. nodosus (VDN LAMP) was developed and evaluated for field use. A sample of 19 sheep flocks (309 sheep) in Victoria, Australia, were tested to determine the optimum conditions for in-field VDN LAMP assay use and sampling, for detecting aprV2-positive D. nodosus infected sheep. VDN LAMP performance was compared to a validated rtPCR that detects aprV2 and the benign strain counterpart, aprB2, using biologically duplicate samples to determine sensitivity and specificity. Flocks were sampled either in winter-spring (moist) or early summer (dry) conditions and had a range of clinical expressions of the disease ovine footrot. Variables considered for optimizing field performance were: sample collection method, sample preparation, clinical expression of disease, and nature of the feet when sampled (moist vs. dry, clean vs. soiled). The test was found to perform best when sheep were sampled with moist, clean feet, using a dry swab with the sample prepared in alkaline polyethylene glycol, pH 13.0, as the collection buffer. A sensitivity of 89% and specificity of 97% was seen when used in-field under these conditions, when compared to aprV2 detection by rtPCR, with "very good" agreement to rtPCR results. This study shows the VDN LAMP test is easy to use in-field to identify the presence of aprV2-positive D. nodosus in sheep flocks.

Please choose payment method:






(PDF emailed within 0-6 h: $19.90)

Accession: 066621151

Download citation: RISBibTeXText

PMID: 30906742

DOI: 10.3389/fvets.2019.00067


Related references

The development and deployment of a field-based loop mediated isothermal amplification assay for virulent Dichelobacter nodosus detection on Australian sheep. Plos one 13(9): E0204310, 2018

Loop-mediated isothermal amplification (LAMP) assay for the diagnosis of fasciolosis in sheep and its application under field conditions. Parasites and Vectors 9: 73, 2016

Development and optimization of a group-specific loop-mediated isothermal amplification (LAMP) assay for the detection of patulin-producing Penicillium species. International Journal of Food Microbiology 298: 20-30, 2019

Development and comparative evaluation of loop mediated isothermal amplification (LAMP) assay for simple visual detection of orf virus in sheep and goats. Molecular and Cellular Probes 29(3): 193-195, 2015

Validation of SYBR green I based closed tube loop mediated isothermal amplification (LAMP) assay and simplified direct-blood-lysis (DBL)-LAMP assay for diagnosis of visceral leishmaniasis (VL). Plos Neglected Tropical Diseases 12(11): E0006922, 2018

Development of reverse transcription-loop-mediated isothermal amplification (RT-LAMP) assay for detection of avian influenza viruses in field specimens. Journal of Veterinary Medical Science 72(4): 519-523, 2010

Detection of Fusarium graminearum DNA using a loop-mediated isothermal amplification (LAMP) assay. International Journal of Food Microbiology 140(2-3): 183-191, 2010

Loop-Mediated Isothermal Amplification (LAMP) for Detection of Phytoplasmas in the Field. Methods in Molecular Biology 1302: 99, 2015

A loop-mediated isothermal amplification (LAMP) assay for the rapid detection of Enterococcus spp. in water. Water Research 122: 62-69, 2017

Development of a loop-mediated isothermal amplification (LAMP) assay for the detection of Anaplasma marginale. Experimental and Applied Acarology 77(1): 65-72, 2019

Optimization of loop-mediated isothermal amplification (LAMP) assays for the detection of Leishmania DNA in human blood samples. Acta Tropica 162: 20-26, 2016

Molecular detection of Coxiella burnetii using an alternative loop-mediated isothermal amplification assay (LAMP). Veterinaria Italiana 51(1): 73-78, 2016

Development of a novel loop-mediated isothermal amplification (LAMP) assay for the detection of Salmonella ser. Enteritidis from egg products. Food Control 88: 190-197, 2018

Development of Loop-Mediated Isothermal Amplification (LAMP) Assay for Rapid Detection of Cannabis sativa. Biological and Pharmaceutical Bulletin 39(7): 1144-1149, 2016

Development of Loop-Mediated Isothermal Amplification (LAMP) Assay for the Rapid Detection of Alternaria alternata. Journal of Aoac International 100(1): 99, 2017