Development of a real-time PCR assay for the detection of the golden mussel ( Limnoperna fortunei , Mytilidae) in environmental samples.

Marcio R Pie,Michelle Z Tadra-Sfeir,Patrícia R Ströher,Antonio Ostrensky,Ricardo Belmonte-Lopes,André O Agostinis

doi:10.1590/0001-3765201720160723

Marcio R Pie, Michelle Z Tadra-Sfeir + Show 4 more

Open Access

https://doi.org/10.1590/0001-3765201720160723

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Abstract

The golden mussel, Limnoperna fortunei, is among the most devastating invasive species in freshwater habitats worldwide, leading to severe environmental disturbances and economic losses. Therefore, management efforts would be greatly improved by methods that efficiently detect and quantify the abundance of the golden mussel in freshwater habitats, particularly in early stages of colonization. In this study, we describe a highly-sensitive real-time PCR assay targeting a 100-bp region of the COI mitochondrial gene of the golden mussel. The method was able to detect as little as 0.225 pg of target DNA. This assay represents an important contribution to surveillance methods, as well as to optimize field measures to contain and manage populations of the golden mussel in its introduced range.

Highlights

The golden mussel Limnoperna fortunei (Mytilidae, Bivalvia) is an invasive bivalve from Southeast Asia rivers (Xu et al 2015)
Recent studies have sought to detect free DNA molecules in suspension, known as eDNA, which usually involves smaller fragments (Ficetola et al 2008, Taberlet et al 2012). eDNA methods have been useful in monitoring programs of invasive species, including frogs (Ficetola et al 2008), fish (Darling and Mahon 2011, Keskin 2014) and mudsnails (Goldberg et al 2013)
We downloaded 57 sequences of the cytochrome c oxidase subunit I (COI) mitochondrial gene of the golden mussel available on genbank, those sequenced by Ghabooli et al (2013) and searched for conserved regions that would allow for the amplification of a short fragment that could be used even in the case of highly degraded environmental DNA

Summary

Introduction

The golden mussel Limnoperna fortunei (Mytilidae, Bivalvia) is an invasive bivalve from Southeast Asia rivers (Xu et al 2015). The efficiency of methods for the management and control of the golden mussel depends heavily on the ability of detecting its presence, in the early stages of its colonization. To this end, Pie et al (2006) developed a set of species-specific primers that could be used to detect golden mussel larvae obtained from filtered environmental water. The large fragment that was amplified (~ 300 bp) requires that the template DNA is relatively well preserved, allowing for its detection only in the presence of fresh larval tissue in the samples. Recent studies have sought to detect free DNA molecules in suspension, known as eDNA, which usually involves smaller fragments (Ficetola et al 2008, Taberlet et al 2012). eDNA methods have been useful in monitoring programs of invasive species, including frogs (Ficetola et al 2008), fish (Darling and Mahon 2011, Keskin 2014) and mudsnails (Goldberg et al 2013)

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