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Updated: Apr 20, 2026
Multiplex PCR Assay for Typing of Staphylococcal Cassette Chromosome Mec Types I to V in Methicillin-resistant Staphylococcus aureus
Published on: September 5, 2013
Designing multiplex PCR system of Campylobacter jejuni for efficient typing by improving monoplex PCR binary typing
Kazuhiro Yamada1, Ami Ibata2, Masahiro Suzuki2
1Department of Microbiology and Medical Zoology, Aichi Prefectural Institute of Public Health, Nagoya 462-8576, Japan; Department of Biological Chemistry, College of Bioscience and Biotechnology, Chubu University, Kasugai, Aichi 487-8501, Japan.
Abstract:
Campylobacter jejuni is responsible for the majority of Campylobacter infections. As the molecular epidemiological study of outbreaks, pulsed-field gel electrophoresis (PFGE) is performed in general. But PFGE has several problems. PCR binary typing (P-BIT) method is a typing method for Campylobacter spp. that was recently developed, and was reported to have a similar discriminatory power and stability to those of PFGE. We modified the P-BIT method from 18 monoplex PCRs to two multiplex PCR systems (mP-BIT). The same results were obtained from monoplex PCRs using original primers and multiplex PCR in the representative isolates. The mP-BIT can analyze 48 strains at a time by using 96-well PCR systems and can identify C. jejuni because mP-BIT includes C. jejuni marker. The typing of the isolates by the mP-BIT and PFGE demonstrated generally concordant results and the mP-BIT method (D = 0.980) has a similar discriminatory power to that of PFGE with SmaI digest (D = 0.975) or KpnI digest (D = 0.987) as with original article. The mP-BIT method is quick, simple and easy, and comes to be able to perform it at low cost by having become a multiplex PCR system. Therefore, the mP-BIT method with two multiplex PCR systems has high potential for a rapid first-line surveillance typing assay of C. jejuni and can be used for routine surveillance and outbreak investigations of C. jejuni in the future.
Insights
A new multiplex PCR binary typing (mP-BIT) method offers a rapid, cost-effective alternative to PFGE for Campylobacter jejuni surveillance. This method provides similar discriminatory power and stability for outbreak investigations.
Area of Science:
- Microbiology
- Molecular Epidemiology
- Infectious Disease Surveillance
Background:
- Campylobacter jejuni causes most Campylobacter infections.
- Pulsed-field gel electrophoresis (PFGE) is standard for molecular epidemiology but has limitations.
- Previous PCR binary typing (P-BIT) showed promise but required optimization.
Purpose of the Study:
- To modify and evaluate a multiplex PCR binary typing (mP-BIT) method for Campylobacter spp.
- To assess mP-BIT's efficiency, discriminatory power, and stability compared to PFGE.
- To establish mP-BIT as a rapid surveillance tool for C. jejuni.
Main Methods:
- Modified the P-BIT method from monoplex to two multiplex PCR systems (mP-BIT).
- Analyzed representative isolates using both original primers and multiplex PCR.
- Compared mP-BIT results with PFGE typing (SmaI and KpnI digests).
Main Results:
- Multiplex PCR (mP-BIT) yielded identical results to monoplex PCR.
- mP-BIT can analyze 48 strains simultaneously using 96-well PCR systems.
- mP-BIT demonstrated high discriminatory power (D = 0.980), comparable to PFGE (D = 0.975–0.987).
- Typing results from mP-BIT and PFGE were generally concordant.
Conclusions:
- The mP-BIT method is quick, simple, easy, and cost-effective.
- mP-BIT is suitable for identifying C. jejuni and analyzing multiple strains efficiently.
- mP-BIT shows high potential as a rapid first-line surveillance and outbreak investigation assay for C. jejuni.
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