Evaluation of a Nested Polymerase Chain Reaction Assay to Differentiate Between Two Genotypes of <i>Porcine Circovirus-2</i>

Lyoo, Kwang‐Soo; Kim, Hyeun Bum; Joo, Heesoo

doi:10.1177/104063870802000304

Cited by 21 publications

(15 citation statements)

References 14 publications

(45 reference statements)

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“…Generally, the nucleotide sequencing has been used to detect PCV2a and PCV2b [12], other approaches included PCR [16], nested PCR [17], PCR-RFLP [18] and TaqMan-based assay [27]. However, these methods were usually time consuming and high technical requirement, and were difficult to popularize in veterinary clinic practice.…”

Section: Discussionmentioning

confidence: 99%

“…These controversial PCV2 new genotypes need further investigation. At present, PCR [16], nested PCR [17], PCR-RFLP [18], TaqMan-based assay [19], nucleotide sequencing and phylogenetic analysis [12] have been used to detect and differentiate PCV2a and PCV2b. However, the application of these methods is limited somewhat by the expensive PCR equipment, the long detection period or the high technical requirements [20,21].…”

Section: Introductionmentioning

confidence: 99%

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Development of a loop-mediated isothermal amplification method to rapidly detect porcine circovirus genotypes 2a and 2b

Qiu

Tian

Zhang

et al. 2012

Virol J

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BackgroundPorcine circovirus type 2 (PCV2), is nowadays associated with a number of diseases known as porcine circovirus-associated diseases (PCVAD), especially postweaning multisystemic wasting syndrome (PMWS). The epidemiological investigation of PCV2 infection was usually conducted by PCR, nested PCR, PCR-RFLP, TaqMan-based assay and nucleotide sequencing. However, there is still no rapid, sensitive and practical method for detecting PCV2 genotypes. As a novel nucleic acid amplification method, the loop-mediated isothermal amplification method (LAMP) has been used to detect a variety of pathogenic microorganisms.ResultsHerein, a LAMP method is developed to detect the genotypes of PCV2. The diagnostic sensitivity of LAMP is 1 copy/reaction for differentiating genotypes PCV2a and PCV2b. The reaction process was completed at 65°C for 1 hour in a water bath. Cross-reactivity assay shows that this method is specific for PCV2a and PCV2b and no reactive for PCV2c and other swine-origin viruses (i.e. CSFV, PRRSV, BVDV, TGEV and PEDV, etc). Identity between LAMP and nested PCR was 92.3% on 52 field clinical samples.ConclusionsLAMP method provides a rapid, sensitive, reliable way to detect PCV2a and PCV2b, and a better means for the large scale investigation of PCV2a and PCV2b infection.

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Development of a loop-mediated isothermal amplification method to rapidly detect porcine circovirus genotypes 2a and 2b

Qiu

Tian

Zhang

et al. 2012

Virol J

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“…The PCV2c genotype has only been found in Denmark to date [31]. In recent years, several reports have suggested that PCV2b is the predominant genotype in many countries worldwide, including Brazil [8,11], Canada [14,15], China [17,21,38], Cuba [29], Denmark [12], Germany [30], Japan [32], Korea [1,2,19,20], Slovakia [37], Spain [10], Sweden [33], Switzerland [39], Thailand [18] and the U.S.A. [6,23]. PCV2b might be more pathogenic than PCV2a [16,32].…”

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confidence: 99%

Genotypic Change and Phylogenetic Analysis of Porcine Circovirus Type 2 in Taiwanese Pig Herds

et al. 2012

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ABSTRACT. Porcine circovirus type 2 (PCV2) is the primary causative agent of postweaning multisystemic wasting syndrome. Two major PCV2 genotypes, PCV2a and PCV2b, have been identified. To explore the prevalence of different subgroups of PCV2 in Taiwan Porcine circovirus type 2 (PCV2) is the primary causative agent of porcine circovirus-associated diseases (PCVADs) in pigs and causes significant economic losses in most pig-producing countries [27]. PCVADs can be subclinical or can include 1 or more clinical manifestations such as multisystemic disease with weight loss, respiratory signs, enteric signs, porcine dermatitis and nephropathy syndrome, and reproductive problems individually or in combination in pigs raised on farms or in other groups of pigs [27]. PCVADs are multifactorial syndromes, and factors that are currently thought to influence the outcome of PCV2 infection include the viral strain, the host, coinfections, and immune modulation [27].PCV2 is a small, non-enveloped, circular, single-stranded DNA virus containing a circular genome of 1766-1768 nucleotides [17,34]. PCV2 is a member of the genus Circovirus of the family Circoviridae [27]. Another member of this genus includes PCV1, which does not produce disease in pigs [35]. The PCV2 genome contains three open reading frames (ORFs). ORF1 encodes two replicase proteins, Rep and Rep' [4]. The viral capsid protein is encoded by ORF2 [25], which is involved in the host immune response [24]. The third ORF, ORF3, is in a different reading frame embedded within ORF1 and encodes a protein associated with apoptosis [22].PCV2 viruses are further classified into three genotypes (2a, 2b, and 2c) that differ with respect to the ORF2 sequence [9,26]. The signature motif (amino acid positions 86-89) of ORF2 can be used to distinguish PCV2a and 2b, which contain the TNKI and SNPR (L) motifs, respectively [6]. The PCV2c genotype has only been found in Denmark to date [31]. In recent years, several reports have suggested that PCV2b is the predominant genotype in many countries worldwide, including Brazil [8,11], Canada [14,15] [16,32]. In Taiwan, the first PCV2 outbreak was confirmed in 2001 [3], and a high prevalence of PCV2 infection was noted thereafter. Up to 92% of pig herds and 68.8% of weaned pigs were infected by PCV2 in Taiwan [7]; however, the genotype prevalence of PCV2 in Taiwan is not currently known. The aim of the present study was to investigate the prevalence of the PCV2 genotype in Taiwan in the last decade. MATERIALS AND METHODS Specimen collection and histopathological examination:Clinical samples (lungs, lymph nodes, spleen, brain and serum) were collected from 785 pigs from 186 herds in central and southern Taiwan during 2002-2011. These samples were mainly acquired from weaning and growing pigs with

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“…Recently, there are several excellent methods that can be used to identify the PCV2a and PCV2b; these include polymerase chain reaction (PCR) assay, nested PCR, real-time PCR and restriction fragment length polymorphism (RFLP) [5,14,15,16]. These methods are convenient and allow the testing of large number of clinical samples.…”

Section: Introductionmentioning

confidence: 99%

Retrospective Study of Porcine Circovirus Type 2a and 2b Between 1999 and 2016 in Taiwan

Lee¹

2017

AVS

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Porcine circovirus type 2 (PCV2) is one of the major swine viral diseases and caused significant economic loss to pig producers worldwide, including Taiwan. PCV2 has been considered as the causative agent of postweaning multisystemic wasting syndrome (PMWS) as well as other clinical diseases. All these associated syndromes have been categorized as PCV2 associated diseases (PCVAD). The purpose of this study was to investigate the positive rate and genetic shift of two distinct genotypes of PCV2, which include PCV2a and PCV2b, in Taiwanese pig farms. A total of 1094 specimens originating from pigs between years 1999 and 2016 were analysed. The PCV2a and PCV2b sequences were amplified and distinguished using 1oop-mediated isothermal amplification (LAMP). Results showed that 24.8% (272/1094) pigs were PCV2a positive, 67.3% (737/1094) were PCV2b positive. These results also indicated that PCV2a was the predominant virus between 1999 and 2001, and that PCV2b became the most prevalent virus since 2003.

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Evaluation of a Nested Polymerase Chain Reaction Assay to Differentiate Between Two Genotypes of Porcine Circovirus-2

Cited by 21 publications

References 14 publications

Development of a loop-mediated isothermal amplification method to rapidly detect porcine circovirus genotypes 2a and 2b

Development of a loop-mediated isothermal amplification method to rapidly detect porcine circovirus genotypes 2a and 2b

Genotypic Change and Phylogenetic Analysis of Porcine Circovirus Type 2 in Taiwanese Pig Herds

Retrospective Study of Porcine Circovirus Type 2a and 2b Between 1999 and 2016 in Taiwan

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