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

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.

The effect of tube trap structure on sampling efficacy and accuracy for golden mussel, <i>Limnoperna fortunei</i>

“…Various methods for early detection of golden mussels, such as molecular analysis (Pie et al, 2006, Boeger et al, 2007, Endo et al, 2009, Pie et al, 2017 have recently been developed. Although these methods are very useful (Darrigran et al 2009), it may be preferable to use simple traps in situations where there are few available funds for detecting the mussels.…”

“…In addition, it can be difficult to directly observe these facilities because water cannot be withdrawn easily from irrigation ponds and reservoirs while the irrigation facilities are in use. Molecular biological techniques that have been advanced in recent years (such as those using environmental DNA), are thought to be effective for detecting the presence or absence of invasive aquatic organisms, although these methods are not yet able to provide quantitative data (Pie et al 2017;Xia et al, 2017). In addition, the density of larvae that can be used as an indicator of the abundance of the mussel fluctuates drastically during the reproductive season (e.g.…”

RFLP pattern determination for the invasive bivalves Limnoperna fortunei (Dunker, 1857) and Corbicula fluminea (Muller, 1774)

“…Bivalve larvae become a major problem when it comes to the identification of these organisms due to the difficulty of morphological distinction in microscopy analysis (Ludwig et al, 2014) at the most crucial moment for the management of invasive species during the early stages of colonization (Pie et al, 2017). Generally, optical identification techniques do not work with most bivalve larvae, since the specific characters appear only in the later stages of development or are visible only on a microscopic level (Garland and Zimmer, 2002).…”

“…Água vol. 13 n. 3, e2172 -Taubaté 2018 Molecular methods for detection of golden mussel were proposed using specific primers (Pie et al, 2006b), but they are restricted to binary responses (Pie et al, 2017), whereas technologies employing Real Time PCR (Endo et al, 2009;Pie et al, 2017) perform a quantification of larvae by indirect means, increasing the time and cost of execution. The same applies to the methodology used for the detection of Corbicula sp.…”

An experimental assessment of the distribution of environmental DNA along the water column

“…After extraction, samples were quantified using rtPCR with a hydrolysis probe (TaqMan) targeting the 100 bp fragment previously amplified COI fragment (Pie et al 2017). Each assay was run in a 10 µL final volume, with concentrations as follows: 75 µM each primer, 25 µM probe and 1 X QuantiNova Probe PCR Kit (Qiagen).…”