A fast and accurate molecular method for the detection of larvae of the golden mussel Limnoperna fortunei (Mollusca: Mytilidae) in plankton samples

Pie, Márcio R.; Boeger, Walter A.; Patella, Luciana; Falleiros, Renan M.

doi:10.1093/mollus/eyi070

Cited by 40 publications

(33 citation statements)

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“…The golden mussel Limnoperna fortunei (Dunker) is an invasive freshwater bivalve species that has been identified as contributing towards various ecological and economic problems (Darrigran et al 1998, Ricciardi 1998, Penchaszadeh et al 2000, Magara et al 2001, Boltovskoy et al 2006, Pie et al 2006. In 2006, this species was designated as an Invasive Alien Species of Japan due to its adverse effects on ecosystems.…”

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confidence: 99%

Molecular based method for the detection and quantification of larvae of the golden mussel Limnoperna fortunei using real-time PCR

Endo

Sato²,

Nogata

2009

Plankton Benthos Res

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A molecular based method for the quantitative detection of larvae of the golden mussel Limnoperna fortunei was developed. Partial nucleotide sequences of the mitochondrial cytochrome c oxidase subunit 1 (CO1) gene in several bivalve species including L. fortunei were analyzed. These nucleotide sequences and the CO1 gene for bivalve species found in habitats similar to L. fortunei (obtained from the EMBL/Genbank/DDBJ databases) were aligned. Based on these results, L. fortunei-specific primers were designed for amplification. DNA extracted from several bivalve species including L. fortunei were subjected to PCR using L. fortunei-specific primers, and amplification of the DNA fragment was confirmed only in PCR assays which utilized template DNA from L. fortunei. No DNA fragments were amplified during the PCR of freshwater planktonic sample lacking golden mussel larvae which were collected from Lake Ohshio (Gunma, Japan). Real-time PCR was performed using fluorescent dye (SYBR Green) detection. Threshold cycles correlated well with the template DNA and number of larvae equivalents, and the R-square values of the standard curves were 0.99 and 0.98, respectively. These results suggest that this real-time PCR-based method can be utilized not only for the identification of L. fortunei in a particular habitat, but also for the quantification of larvae of this species in natural environments.

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mentioning

confidence: 99%

Molecular based method for the detection and quantification of larvae of the golden mussel Limnoperna fortunei using real-time PCR

Endo

Sato²,

Nogata

2009

Plankton Benthos Res

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“…The total DNA was extracted using the EZ-DNA kit (Biosystems, Brazil), following the manufacturer's instructions. Adult oysters were identified using the protocol developed by PIE et al (2006a).…”

Section: Methodsmentioning

confidence: 99%

“…The sensitivity of the optimized Multiplex PCR protocol was tested using extracts from adult tissue with 10-10.000 fold dilutions of an initial concentration of 2 ng/µL (0.2, 0.02, 0.002, 0.0002 ng/µL, respectively). A combined test for sensitivity and specificity was performed using the equivalent of the DNA content of a single bivalve larva (28.5 ng according to PIE et al 2006a) added to the full extract of a zooplankton sample considered negative for larvae of both species of Crassostrea (1000 L of water filtered with 100 µm mesh size). Treatments were designed to simulate the absence, the presence of a single larva, and the simultaneous presence of one larva of each target species, submitted to the optimized multiplex PCR protocol.…”

Section: Methodsmentioning

confidence: 99%

“…Fortunately, molecular protocols are being developed to allow simultaneous detection and identification of one or more species of interest in different types of samples, including plankton samples (MORGAN & ROGERS 2001, ANIL et al 2002, KOTTA et al 2006, PIE et al 2006a. Indeed, identification of adult species of Crassostrea by molecular markers is becoming a common procedure (WANG & GUO 2007, PIE et al 2006b, PATIL et al 2005.…”

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confidence: 99%

“…However, the basic limitations that exist today to implementing molecular methods are the financial costs of using them in monitoring studies. Molecular methodologies were developed by PIE et al (2006a) for screening larvae of the golden mussel, Limnoperna fortunei (Dunker, 1857) in zooplankton samples. That method has been shown to be efficient for continuous monitoring in rivers and lakes of the state of Parana, Brazil (BOEGER et al 2007, DARRIGRAN et al 2009, TSCHÁ et al 2009.…”

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confidence: 99%

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A molecular method to detect and identify the native species of southwestern Atlantic Crassostrea (Mollusca: Ostreidae)

Ludwig¹,

Patella²,

Stoiev³

et al. 2011

Zoologia (Curitiba, Impr.)

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Among oysters, species of Crassostrea (Sacco, 1897) are the most attractive to aquaculture. In Brazil, the genus is represented by C. rhizophorae (Guilding, 1828) and C. brasiliana (Lamarck, 1819). Because the maturation and breeding technology is not well developed for these species, aquaculturists need a reliable method to decide the correct time to place spat collectors in the field, and to identify both species, which are morphologically similar. In this study a specific Multiplex PCR protocol was developed, using one pair of universal primers from 18S rDNA as a positive control and a pair of specific primers for each target species. The sensitivity and specificity of the protocol was evaluated. It detected C. rhizophorae DNA in low concentrations, and C. brasiliana DNA in even lower concentrations. Further, the Multiplex PCR proved efficient in detecting DNA in concentrations equivalent to that of a single larva of each species, either separated or combined, when mixed with total DNA extract of a plankton sample representing 1000 L of filtered water. Field tests confirmed the applicability of the protocol, which holds the promise to become an important tool for aquaculture or conservation programs, allowing for the continuous monitoring of the life cycle of C. brasiliana and C. rhizophorae, by detecting the right periods of larval release and settlement

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Strategies and Measures to Prevent Spread of Invasive Species

Darrigran

Damborenéa

2015

Limnoperna Fortunei

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On the basis of the fouling caused by the golden mussel, Limnoperna fortunei, in industrial facilities and power plants (human environment), and its impacts on the ecosystem (natural environment), several strategies and measures for the mitigation of problems and prevention of further spread are discussed. At the local level, monitoring and early detection of the golden mussel can be accomplished through different methods, including those aimed at juveniles and adults, and also those aimed at their planktonic larvae. Priorities for designing bioinvasion management strategies should focus on generation of scientific knowledge, management, and actions at the sociopolitical level. Mitigation measures are closely related to the invasion phase, whereby the cost increases and the probability of eradication decreases over time.

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A fast and accurate molecular method for the detection of larvae of the golden mussel Limnoperna fortunei (Mollusca: Mytilidae) in plankton samples

Cited by 40 publications

References 0 publications

Molecular based method for the detection and quantification of larvae of the golden mussel Limnoperna fortunei using real-time PCR

Molecular based method for the detection and quantification of larvae of the golden mussel Limnoperna fortunei using real-time PCR

A molecular method to detect and identify the native species of southwestern Atlantic Crassostrea (Mollusca: Ostreidae)

Strategies and Measures to Prevent Spread of Invasive Species

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