Laboratory assessment of sensitive molecular tools for detection of low levels of Echinococcus multilocularis-eggs in fox (Vulpes vulpes) faeces

Particle and Fibre Toxicology

Rebecca K Davidson²  Saraya Tavornpanich²  Åsa Hagström¹  Mats Isaksson¹  Øivind Øines² 

[1] National Veterinary Institute, Uppsala 75189, Sweden;Norwegian Veterinary Institute, Post boks 750 Sentrum, 0106, Oslo, Norway

关键词: Replicates;    Molecular detection;    Real-time PCR;    Surveillance tool;    Echinococcus multilocularis;   

Others  :  806140
DOI  :  10.1186/1756-3305-7-246

 received in 2013-12-27, accepted in 2014-05-14,  发布年份 2014

【 摘 要 】

Background

In endemic areas with very low infection prevalence, the frequency and intensity of Echinococcus multilocularis can be extremely low. This necessitates efficient, specific and sensitive molecular tools. We wanted to compare the existing molecular tools, used in the Norwegian national surveillance programme, and compare these with new techniques for detection of this zoonotic pathogen in fox faeces. Here we present the results of screening samples containing a known level of E. multilocularis eggs with two highly sensitive DNA isolation and extraction methods combined with one conventional PCR and three real-time PCR methods for detection.

Methods

We performed a comparison of two extraction protocols; one based on sieving of faecal material and one using targeted DNA sampling. Four methods of molecular detection were tested on E. multilocularis-egg spiked fox faeces.

Results

There were significant differences between the multiplex PCR/egg sieving DNA extraction methods compared to the new DNA fishing method and the three real-time PCR assays. Results also indicate that replicates of the PCR-reactions improve detection sensitivity when egg numbers are low.

Conclusions

The results indicate that the use of real-time PCR combined with targeted DNA extraction, improves the sensitivity of E. multilocularis detection in faecal samples containing low numbers of E. multilocularis eggs. Results also indicate the importance of replicates of the PCR-reactions when pathogen levels are low.

【 授权许可】

   
2014 Øines et al.; licensee BioMed Central Ltd.

【 图 表 】

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