A Rapid Colorimetric Assay for On-Site Authentication of Cephalopod Species

Tatulli, Giuseppina; Cecere, Paola; Maggioni, Davide; Galimberti, Andrea; Pompa, Pier Paolo

doi:10.3390/bios10120190

Cited by 10 publications

(5 citation statements)

References 35 publications

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“…We found a total of 16 COI-haplotypes for L. forbesii, whereof three have been reported before [32][33][34][35][36][37][38][39][40] (H7 and H10, "East Atlantic" clade; H25, Azorean clade, see Fig. 3).…”

Section: Discussionsupporting

confidence: 56%

“…The sequences obtained were analysed with the software "CEQ8000 Genetic Analysis System" (Version 9.0.25, Beckman Coulter GmbH, Germany) and an alignment of 249 sequences was created ("BioEdit" version 7.2.5. 32 ) of which 28 were retrieved from GenBank [32][33][34][35][36][37][38][39][40] . The phylogenetic analyses (Maximum Likelihood, ML) was performed using "MEGA" version 6 41 with 1000 bootstraps.…”

Section: Phylogeography Via Coi-sequencingmentioning

confidence: 99%

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Phylogeography of the veined squid, Loligo forbesii, in European waters

Göpel

Oesterwind²,

Barrett

et al. 2022

Sci Rep

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The veined squid, Loligo forbesii Steenstrup, 1856, occurs at the European Shelf areas including the Azores and represents a valuable resource for the European commercial fishery in the North East Atlantic. However, very little is known about its population structure and phylogeography. This lack of knowledge also impedes the development of sustainable fishery management for this species. The present study combined the use of two types of markers that retrieve patterns of gene flow in different time spans; the analysis of 16 nuclear microsatellites and sequencing of the mitochondrial cytochrome oxidase subunit I (COI). Whereas the high mutation rate of microsatellites allows the description of recent patterns of connectivity in species, the lower mutation rate of COI provides phylogeographic patterns on a longer timescale. A total of 347 individuals of L. forbesii were investigated from nearly the entire distribution range of the species, including the North East Atlantic Shelf, the Azores and the Mediterranean. Individuals from the Western and Eastern Mediterranean Sea have never been included in a genetic study before. We were able to analyse COI sequences from all 12 sampling areas and define three clades of L. forbesii. Due to our large sampling area, we are presenting 13 COI-haplotypes that were previously unknown. The microsatellite analysis does not include the Azores but three main clades could be identified at the remaining 11 sampling sites. Low FST values indicate gene flow over large geographical distances. However, the genetically significant differences and an additional slight grouping in the microsatellite structure reveal that geographical barriers seem to influence the population structure and reduce gene flow. Furthermore, both markers provide strong evidence that the observed phylogeographic pattern reflects the geographical history of the Azores and the Mediterranean Sea.

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Section: Discussionsupporting

confidence: 56%

Section: Phylogeography Via Coi-sequencingmentioning

confidence: 99%

Phylogeography of the veined squid, Loligo forbesii, in European waters

Göpel

Oesterwind²,

Barrett

et al. 2022

Sci Rep

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show abstract

“…After the optimization of our test, we attempted to develop a simplified and instrument-free approach with a naked-eye readout through a clear color change of the reaction samples. To this end, we exploited the pH-sensitive LAMP strategy using the Cresol Red dye, which results in a pink-to-yellow color change upon target amplification, enabling the identification of positive samples by simple visual inspection [ 44 , 45 , 46 , 47 ].…”

Section: Resultsmentioning

confidence: 99%

“…Once it was assessed that the colorimetric procedure was able to consistently detect the presence of C. tyrobutyricum spores even at a low concentration, we investigated its implementation in milk samples using artificially contaminated specimens and the same extraction procedure described above with no purification steps. In these tests, we took advantage of the high tolerance of the LAMP reaction to matrix contaminants [ 45 ]. As displayed in Figure 5 , the experimental results showed the achievement of a LoD of 10 spores/reaction, with a 50% detection reliability for 5 spores/reaction.…”

Section: Resultsmentioning

confidence: 99%

Colorimetric Point-of-Care Detection of Clostridium tyrobutyricum Spores in Milk Samples

et al. 2021

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Clostridium tyrobutyricum represents the main spoiling agent responsible for late blowing defects (LBD) in hard and semi-hard cheeses. Its spores are resistant to manufacturing procedures and can germinate during the long ripening process, causing the burst of the cheese paste with a consequent undesirable taste. The lower quality of blown cheeses leads to considerable financial losses for the producers. The early identification of spore contaminations in raw milk samples thus assumes a pivotal role in industrial quality control. Herein, we developed a point of care (POC) testing method for the sensitive detection of C. tyrobutyricum in milk samples, combining fast DNA extraction (with no purification steps) with a robust colorimetric loop-mediated isothermal amplification (LAMP) technique. Our approach allows for the sensitive and specific detection of C. tyrobutyricum spores (limit of detection, LoD: ~2 spores/mL), with the advantage of a clear naked-eye visualization of the results and a potential semi-quantitative discrimination of the contamination level. In addition, we demonstrated the feasibility of this strategy using a portable battery-operated device that allowed both DNA extraction and amplification steps, proving its potential for on-site quality control applications without the requirement of sophisticated instrumentation and trained personnel.

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“…phenol red and HNB. Since then, the assay has served as a springboard for various other assays that address the diagnostic needs in a wide range of emerging point-of-need applications, predominantly those in aquaculture [ 17 , 36 ] where demand for simple, robust one-step test kits is high.…”

Section: Discussionmentioning

confidence: 99%