Dataset Information

A simple and rapid method for detection of Goose Parvovirus in the field by loop-mediated isothermal amplification.

ABSTRACT: BACKGROUND: Goose parvovirus (GPV) is a Dependovirus associated with latent infection and mortality in geese. Currently, in a worldwide scale, GPV severely affects geese production. The objective of this study is to develop a loop-mediated isothermal amplification (LAMP) method for the sensitive, rapid, and inexpensive detection of GPV in the field. RESULTS: A set of six specific primers was designed by targeting the GPV VP3 DNA. With Bst DNA polymerase large fragment, the target DNA could be amplified at 65 degrees C as early as 20 min of incubation in a simple water bath. A positive reaction was identified through the detection of the LAMP product by color change visible to the naked eye. The detection limit of the assay was 28 copies/microl of plasmid pVP3, and with equal sensitivity and specificity to fluorescent quantitative real-time PCR (FQ-PCR). CONCLUSIONS: The high sensitivity, specificity, and simplicity, as well as the high throughput, make this method suitable for specific detection of GPV infection in both field conditions and laboratory settings. The utilization of complicated equipment and conduct of technical training on the GPV LAMP were not necessary.

SUBMITTER: Yang J

PROVIDER: S-EPMC2829533 | biostudies-literature | 2010

REPOSITORIES: biostudies-literature

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A simple and rapid method for detection of Goose Parvovirus in the field by loop-mediated isothermal amplification.

Yang JinLong J Yang Rui R Cheng AnChun A Wang MingShu M Fu LiZhi L Yang SongQuan S Zhang SuHui S Yang Liu L Xu ZhiYong Z

Virology journal 20100121

<h4>Background</h4>Goose parvovirus (GPV) is a Dependovirus associated with latent infection and mortality in geese. Currently, in a worldwide scale, GPV severely affects geese production. The objective of this study is to develop a loop-mediated isothermal amplification (LAMP) method for the sensitive, rapid, and inexpensive detection of GPV in the field.<h4>Results</h4>A set of six specific primers was designed by targeting the GPV VP3 DNA. With Bst DNA polymerase large fragment, the target DN ...[more]

PMID: 20092637

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Project description:Bacillus Calmette-Guérin (BCG) is widely used as a live attenuated vaccine against Mycobacterium tuberculosis and is an agent for standard prophylaxis against the recurrence of bladder cancer. Unfortunately, it can cause severe infectious diseases, especially in immunocompromised patients, and the ability to immediately distinguish BCG from other M. tuberculosis complexes is therefore important. In this study, we developed a simple and easy-to-perform identification procedure using loop-mediated amplification (LAMP) to detect deletions within the region of difference, which is deleted specifically in all M. bovis BCG strains. Reactions were performed at 64 °C for 30 min and successful targeted gene amplifications were detected by real-time turbidity using a turbidimeter and visual inspection of color change. The assay had an equivalent detection limit of 1.0 pg of genomic DNA using a turbidimeter whereas it was 10 pg with visual inspection, and it showed specificity against 49 strains of 44 pathogens, including M. tuberculosis complex. The expected LAMP products were confirmed through identical melting curves in real-time LAMP procedures. We employed the Procedure for Ultra Rapid Extraction (PURE) kit to isolate mycobacterial DNA and found that the highest sensitivity limit with a minimum total cell count of mycobacterium (including DNA purification with PURE) was up to 1 × 10(3) cells/reaction, based on color changes under natural light with FDA reagents. The detection limit of this procedure when applied to artificial serum, urine, cerebrospinal fluid, and bronchoalveolar lavage fluid samples was also about 1 × 10(3) cells/reaction. Therefore, this substitute method using conventional culture or clinical specimens followed by LAMP combined with PURE could be a powerful tool to enable the rapid identification of M. bovis BCG as point-of-care testing. It is suitable for practical use not only in resource-limited situations, but also in any clinical situation involving immunocompromised patients because of its convenience, rapidity, and cost effectiveness.

Dataset Information

A simple and rapid method for detection of Goose Parvovirus in the field by loop-mediated isothermal amplification.

Publications

A simple and rapid method for detection of Goose Parvovirus in the field by loop-mediated isothermal amplification.

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