Genomic sequencing of a frog virus 3 strain from cultured American bullfrogs (Lithobates catesbeianus) in Brazil

Ferreira, Cláudia Maris; Subramaniam, Kuttichantran; Sousa, Ricardo Luiz Moro de; Tavares, Loiane Sampaio; Corrêa, Thaís Camilo; Waltzek, Thomas B.

doi:10.1007/s00705-021-05094-y

Cited by 5 publications

(3 citation statements)

References 15 publications

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“…In particular, FV3 infects not only amphibians but also fishes (Price et al 2017) and reptiles (Waltzek et al 2014), but there have been no such reports in fishes and reptiles in South Korea (Do et al 2005;Kim et al 2013;Lee et al 2019). FV3 has been detected in various wild and cultured amphibian populations throughout the United States and Canada (Duffus et al 2015;Price et al 2017) and from introduced American bullfrogs in Brazil, the United Kingdom, Japan, and Taiwan (Ferreira et al 2021;Gray et al 2007;Hsieh et al 2021;Une et al 2009). Our results show that FV3-like ranaviruses in the genus Ranavirus could be the dominant ranavirus species infecting anuran amphibians in South Korea.…”

Section: Resultsmentioning

confidence: 54%

Comparison of detective ranavirus with major capsid protein gene from infected frogs (Pelophylax nigromaculatus and Lithobates catesbeianus) in South Korea

et al. 2022

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Ranaviruses are a primary cause of amphibian extinctions. More consistent ranavirus-infection reports and genetic characterizations of identified viruses are urgently needed, particularly from Asian countries. The objectives of this study were to obtain the partial major capsid protein (MCP) gene sequences (506 bp) of the ranavirus responsible for infecting frogs in South Korea, as our previous research had confirmed using qPCR, and to evaluate their genetic relationships with other previously reported ranavirus sequences. Three different ranavirus MCP sequences were obtained from Pelophylax nigromaculatus and Lithobates catesbeianus. All six different types of MCP sequence from the ranavirus identified in South Korea to date belonged to the Frog virus 3 (FV3)-like virus group in the genus Ranavirus. To better understand the origin and spread of ranaviruses in South Korea, further infection reports and full genome analyses of the identified ranaviruses are needed.

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Section: Resultsmentioning

confidence: 54%

Comparison of detective ranavirus with major capsid protein gene from infected frogs (Pelophylax nigromaculatus and Lithobates catesbeianus) in South Korea

et al. 2022

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“…The development of detection methods for FV3 has been a stagnant issue for many years. However, FV3 is becoming a global threat to biodiversity and aquaculture. − In this study, an RPA-CRISPR/Cas12-AI system is established, optimized, and evaluated, providing a novel POC detection system for FV3. Furthermore, because crRNA is reprogrammable, the integrated system can be used for detecting any viral DNA target.…”

Section: Discussionmentioning

confidence: 99%

Detection of Frog Virus 3 by Integrating RPA-CRISPR/Cas12a-SPM with Deep Learning

Lei,

Lian,

Zhang

et al. 2023

ACS Omega

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A fast, easy-to-implement, highly sensitive, and point-of-care (POC) detection system for frog virus 3 (FV3) is proposed. Combining recombinase polymerase amplification (RPA) and CRISPR/Cas12a, a limit of detection (LoD) of 100 aM (60.2 copies/μL) is achieved by optimizing RPA primers and CRISPR RNAs (crRNAs). For POC detection, smartphone microscopy is implemented, and an LoD of 10 aM is achieved in 40 min. The proposed system detects four positive animal-derived samples with a quantitation cycle (Cq) value of quantitative PCR (qPCR) in the range of 13 to 32. In addition, deep learning models are deployed for binary classification (positive or negative samples) and multiclass classification (different concentrations of FV3 and negative samples), achieving 100 and 98.75% accuracy, respectively. Without temperature regulation and expensive equipment, the proposed RPA-CRISPR/Cas12a combined with smartphone readouts and artificial-intelligence-assisted classification showcases the great potential for FV3 detection, specifically POC detection of DNA virus.

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“…The development of detection methods for FV3 has been stagnant for many years. On the other hand, FV3 is becoming a global threat to biodiversity and aquaculture (Ferreira et al 2021; Cozad et al 2020; Oliveira et al 2019). In this study, we establish, optimize, and evaluate the RPA-CRISPR/Cas12-AI system, providing a novel POC detection system for FV3.…”

Section: Discussionmentioning

confidence: 99%

Detection of Frog virus 3 via the system integrating RPA-CRISPR/Cas12a-SPM with deep learning

Lei

Lian

Zhang

et al. 2022

Preprint

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Frog virus 3 (FV3, genera Ranavirus, family Iridoviridae), a double-stranded DNA virus, results in irreparable damage to biodiversity and significant economic losses to aquaculture. Although the existing FV3 detection methods are of high sensitivity and specificity, the complex procedure and requirement of expensive instruments limit their practical implantation. Herein, we develop a fast, easy-to-implement, highly sensitive, and point-of-care (POC) detection system for FV3. Combining recombinase polymerase amplification (RPA) and CRISPR/Cas12a, we achieve a limit of detection (LoD) of 100 aM (60.2 copies/μL) by optimizing RPA primers and CRISPR RNAs (crRNAs). For POC detection, we build a smartphone microscopy (SPM) and achieve an LoD of 10 aM within 40 minutes. Four positive animal-derived samples with a quantitation cycle (Cq) value of quantitative PCR (qPCR) in the range of 13 to 32 are detectable by the proposed system. In addition, we deploy deep learning models for binary classification (positive or negative samples) and multiclass classification (different concentrations of FV3 and negative samples), achieving 100% and 98.75% accuracy, respectively. Without temperature regulation and expensive equipment, RPA-CRISPR/Cas12a combined with a smartphone readout and artificial intelligence (AI) assisted classification shows great potential for FV3 detection. This integrated system holds great promise for POC detection of aquatic DNA pathogens.

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Genomic sequencing of a frog virus 3 strain from cultured American bullfrogs (Lithobates catesbeianus) in Brazil

Cited by 5 publications

References 15 publications

Comparison of detective ranavirus with major capsid protein gene from infected frogs (Pelophylax nigromaculatus and Lithobates catesbeianus) in South Korea

Comparison of detective ranavirus with major capsid protein gene from infected frogs (Pelophylax nigromaculatus and Lithobates catesbeianus) in South Korea

Detection of Frog Virus 3 by Integrating RPA-CRISPR/Cas12a-SPM with Deep Learning

Detection of Frog virus 3 via the system integrating RPA-CRISPR/Cas12a-SPM with deep learning

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