scholarly journals A Multiplex PCR Assay Combined with Capillary Electrophoresis for the Simultaneous Identification of Atlantic Cod, Pacific Cod, Blue Whiting, Haddock, and Alaska Pollock

Foods ◽  
2021 ◽  
Vol 10 (11) ◽  
pp. 2631
Author(s):  
Yu-Min Lee ◽  
Shinyoung Lee ◽  
Hae-Yeong Kim
Keyword(s):  
Capillary Electrophoresis ◽  
Multiplex Pcr ◽  
Atlantic Cod ◽  
Limit Of Detection ◽  
Pacific Cod ◽  
Pcr Assay ◽  
Food Fraud ◽  
Blue Whiting ◽  
Multiplex Pcr Assay ◽  
Mitochondrial Cytochrome B

With an increased consumption of seafood products, food fraud with fish resources has been continuously reported. In particular, codfish has been exploited worldwide as a processed product in fresh, frozen, smoked, canned, or ready-to-eat dish forms. However, it is challenging to identify processed fish products after processing because of their similar morphological characteristics. Substitution and mislabeling of codfish among different species are also happening deliberately or unintentionally. Thus, it is necessary to distinguish cod species to prevent fish adulteration and food fraud. In this study, we developed a multiplex PCR for simultaneously identifying five cod species within Gadidae using capillary electrophoresis. Then, their species-specific primer sets were designed by targeting the mitochondrial cytochrome b gene. Subsequently, the amplicon sizes obtained were 237 bp, 204 bp, 164 bp, 138 bp, and 98 bp for Atlantic cod, Pacific cod, blue whiting, haddock, and Alaska pollock, respectively. The specificity of each primer was further tested using 19 fish species, and no cross-reactivity was observed. The limit of detection of this multiplex PCR assay was 1 pg. The developed multiplex PCR assay can be applied to 40 commercial food products successfully. This detection method will be efficient for managing seafood authentication by simultaneously analyzing multiple cod species.

scholarly journals Single-Step Multiplex PCR Assay for Determining 92 Pneumococcal Serotypes

2016 ◽  
Vol 54 (8) ◽  
pp. 2197-2200 ◽  
Author(s):  
José M. Marimón ◽  
María Ercibengoa ◽  
Erica Santacatterina ◽  
Marta Alonso ◽  
Emilio Pérez-Trallero
Keyword(s):  
Capillary Electrophoresis ◽  
Multiplex Pcr ◽  
Disease Surveillance ◽  
Pneumococcal Disease ◽  
Cost Effective ◽  
Single Step ◽  
Pneumococcal Serotypes ◽  
Pcr Assay ◽  
Single Tube ◽  
Multiplex Pcr Assay

For pneumococcal disease surveillance, simple and cost-effective methods capable of determining all serotypes are needed. Combining a single-tube multiplex PCR with fluorescently labeled primers followed by amplicon analysis using automated fluorescent capillary electrophoresis, each serotype of 92 reference isolates and 297 recently collected clinical isolates was successfully determined.

scholarly journals Species Identification of Red Deer (Cervus elaphus), Roe Deer (Capreolus capreolus), and Water Deer (Hydropotes inermis) Using Capillary Electrophoresis-Based Multiplex PCR

Foods ◽  
2020 ◽  
Vol 9 (8) ◽  
pp. 982
Author(s):  
Mi-Ju Kim ◽  
Yu-Min Lee ◽  
Seung-Man Suh ◽  
Hae-Yeong Kim
Keyword(s):  
Capillary Electrophoresis ◽  
Multiplex Pcr ◽  
Cervus Elaphus ◽  
Red Deer ◽  
Roe Deer ◽  
Capreolus Capreolus ◽  
Target Species ◽  
Pcr Assay ◽  
Multiple Targets ◽  
Multiplex Pcr Assay

To provide consumers correct information on meat species, specific and sensitive detection methods are needed. Thus, we developed a capillary electrophoresis-based multiplex PCR assay to simultaneously detect red deer (Cervus elaphus), roe deer (Capreolus capreolus), and water deer (Hydropotes inermis). Specific primer sets for these three species were newly designed. Each primer set only amplified target species without any reactivity against non-target species. To identify multiple targets in a single reaction, multiplex PCR was optimized and combined with capillary electrophoresis to increase resolution and accuracy for the detection of multiple targets. The detection levels of this assay were 0.1 pg for red deer and roe deer and 1 pg for water deer. In addition, its applicability was demonstrated using various concentrations of meat DNA mixtures. Consequently, as low as 0.1% of the target species was detectable using the developed method. This capillary electrophoresis-based multiplex PCR assay for simultaneous detection of three types of deer meat could authenticate deer species labeled on products, thus protecting consumers from meat adulteration.

scholarly journals Detection of porcine circoviruses in clinical specimens using multiplex PCR in Hubei, central China

2018 ◽  
Author(s):  
Keli Yang ◽  
Zuwu Jiao ◽  
Danna Zhou ◽  
Rui Guo ◽  
Zhengying Duan ◽  
...  
Keyword(s):  
Multiplex Pcr ◽  
Infection Rate ◽  
Target Genes ◽  
Limit Of Detection ◽  
Central China ◽  
Clinical Samples ◽  
Monitoring And Control ◽  
Pcr Assay ◽  
Tissue Samples ◽  
Multiplex Pcr Assay

In order to detect and simultaneously discriminate PCV1, PCV2 and PCV3, a multiplex PCR assay was developed and used to detect clinical samples in this study. Each of target genes of PCV1, PCV2 and PCV3 was amplified using the designed primers, while no other porcine viruses genes were detected. The limit of detection of the assay was 10 copies/μL of PCV1, PCV2 and PCV3. The tissue samples from eight pig farms were detected using the multiplex PCR assay. The results showed that PCV1, PCV2 and PCV3 are co-circulating in central China. The PCV1, PCV2 and PCV3 singular infection rate was 52.4% (150/286), 61.2% (175/286) and 45.1% (129/286), respectively, while the PCV1 and PCV2 co-infection rate was 11.2% (32/286), the PCV1 and PCV3 co-infection rate was 5.9% (17/286), the PCV2 and PCV3 co-infection rate was 23.4% (67/286), and the PCV1, PCV2 and PCV3 co-infection rate was 1.7% (5/286), respectively, which were 100% consistent with the sequencing method and Real-time PCR methods. It proved that this multiplex PCR assay could be used as a differential diagnostic tool for monitoring and control of PCVs in the field. The results also indicate that the PCVs infection and their co-infection are severe in Hubei Province, Central China.

Design of a multiplex PCR assay for the simultaneous detection and confirmation of Neisseria gonorrhoeae

2010 ◽  
Vol 63 (5) ◽  
pp. 431-433 ◽  
Author(s):  
Isabelle O'Callaghan ◽  
Daniel Corcoran ◽  
Brigid Lucey
Keyword(s):  
Neisseria Gonorrhoeae ◽  
Multiplex Pcr ◽  
Internal Control ◽  
Limit Of Detection ◽  
Simultaneous Detection ◽  
Pcr Primers ◽  
Real Potential ◽  
Pcr Assay ◽  
Neisseria Species ◽  
Multiplex Pcr Assay

ObjectivesTo improve the detection of Neisseria gonorrhoeae by designing a multiplex PCR assay using two N gonorrhoeae-specific genes as targets, thereby providing detection and confirmation of a positive result simultaneously.MethodsPCR primers were designed to detect two N gonorrhoeae genes, namely porA and pgi1. Primers for an internal control targeting the ompW gene of Vibrio cholerae were also designed and incorporated in the assay. The DNA of 45 clinical isolates including 33 N gonorrhoeae isolates, seven non-gonococcal Neisseria species, and five non-Neisseria species was tested using the multiplex PCR assay.ResultsAll 33 N gonorrhoeae isolates were successfully detected by the assay and none of the non-gonococcal isolates was detected. The assay showed a sensitivity and specificity of 100%, and a limit of detection of 1.25 ng of DNA.ConclusionThis multiplex PCR assay offers a sensitive and specific assay suitable for the detection of N gonorrhoeae, and offers real potential for diagnostic use.

scholarly journals Detection of porcine circoviruses in clinical specimens using multiplex PCR in Hubei, central China

2018 ◽  
Author(s):  
Keli Yang ◽  
Zuwu Jiao ◽  
Danna Zhou ◽  
Rui Guo ◽  
Zhengying Duan ◽  
...  
Keyword(s):  
Multiplex Pcr ◽  
Infection Rate ◽  
Target Genes ◽  
Limit Of Detection ◽  
Central China ◽  
Clinical Samples ◽  
Monitoring And Control ◽  
Pcr Assay ◽  
Tissue Samples ◽  
Multiplex Pcr Assay

In order to detect and simultaneously discriminate PCV1, PCV2 and PCV3, a multiplex PCR assay was developed and used to detect clinical samples in this study. Each of target genes of PCV1, PCV2 and PCV3 was amplified using the designed primers, while no other porcine viruses genes were detected. The limit of detection of the assay was 10 copies/μL of PCV1, PCV2 and PCV3. The tissue samples from eight pig farms were detected using the multiplex PCR assay. The results showed that PCV1, PCV2 and PCV3 are co-circulating in central China. The PCV1, PCV2 and PCV3 singular infection rate was 52.4% (150/286), 61.2% (175/286) and 45.1% (129/286), respectively, while the PCV1 and PCV2 co-infection rate was 11.2% (32/286), the PCV1 and PCV3 co-infection rate was 5.9% (17/286), the PCV2 and PCV3 co-infection rate was 23.4% (67/286), and the PCV1, PCV2 and PCV3 co-infection rate was 1.7% (5/286), respectively, which were 100% consistent with the sequencing method and Real-time PCR methods. It proved that this multiplex PCR assay could be used as a differential diagnostic tool for monitoring and control of PCVs in the field. The results also indicate that the PCVs infection and their co-infection are severe in Hubei Province, Central China.

Usefulness of a Multiplex PCR for Detection of Diarrheagenic Escherichia coli in a Diagnostic Microbiology Laboratory Setting

2016 ◽  
Vol 1 (2) ◽  
pp. 38-42 ◽  
Author(s):  
Khairun Nessa ◽  
Dilruba Ahmed ◽  
Johirul Islam ◽  
FM Lutful Kabir ◽  
M Anowar Hossain
Keyword(s):  
Escherichia Coli ◽  
Multiplex Pcr ◽  
Clinical Laboratory ◽  
Proteinase K ◽  
Microbiology Laboratory ◽  
Pcr Assay ◽  
E Coli ◽  
Diarrheagenic Escherichia Coli ◽  
Multiplex Pcr Assay ◽  
Diagnostic Microbiology

A multiplex PCR assay was evaluated for diagnosis of diarrheagenic Escherichia coli in stool samples of patients with diarrhoea submitted to a diagnostic microbiology laboratory. Two procedures of DNA template preparationproteinase K buffer method and the boiling method were evaluated to examine isolates of E. coli from 150 selected diarrhoeal cases. By proteinase K buffer method, 119 strains (79.3%) of E. coli were characterized to various categories by their genes that included 55.5% enteroaggregative E. coli (EAEC), 18.5% enterotoxigenic E. coli (ETEC), 1.7% enteropathogenic E. coli (EPEC), and 0.8% Shiga toxin-producing E. coli (STEC). Although boiling method was less time consuming (<24 hrs) and less costly (<8.0 US $/ per test) but was less efficient in typing E. coli compared to proteinase K method (41.3% vs. 79.3% ; p<0.001). The sensitivity and specificity of boiling method compared to proteinase K method was 48.7% and 87.1% while the positive and negative predictive value was 93.5% and 30.7%, respectively. The majority of pathogenic E. coli were detected in children (78.0%) under five years age with 53.3% under one year, and 68.7% of the children were male. Children under 5 years age were frequently infected with EAEC (71.6%) compared to ETEC (24.3%), EPEC (2.7%) and STEC (1.4%). The multiplex PCR assay could be effectively used as a rapid diagnostic tool for characterization of diarrheagenic E. coli using a single reaction tube in the clinical laboratory setting.Bangladesh J Med Microbiol 2007; 01 (02): 38-42

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