Infect Chemother.  2010 Aug;42(4):237-240. 10.3947/ic.2010.42.4.237.

Further Evaluation of Multiplex PCR for Rapid Detection of Salmonella Typhi

• Lee DY
• Min JE
• Lee E
• Kim SH
• Oh HB
• Park MS

• KMID: 1457384
• DOI: http://doi.org/10.3947/ic.2010.42.4.237

Abstract

Typhoid fever is a class I legally designated communicable disease in Korea; and if remains as an important public health problem in many developing countries. It takes at least 3-5 days to detect and identify Salmonella Typhi (S. Typhi) by classical diagnostic method. For this reason, multiplex PCR (mPCR) was evaluated in detecting and identifying S. Typhi. In this study, forty-three bacterial strains, which consisted of 42 Salmonella enterica serovars and one Citrobacter freundii. were used to evaluate the promptness of mPCR in detecting and identifying S. Typhi. mPCR was performed with four genes which were known for representing Salmonella spp and/or S. Typhi: invA, fliC-d, viaB and prt. invA and prt gene was amplified in all strains and viaB gene was in only S. Typhi. fliC-d gene was amplified in three serovars: S. Typhi, S. Schwarzengrund and S. Livingstone. After specificity test, mPCR was modified as triplex PCR with three genes (invA, fliC-d, and viaB) and the sensitivity test was performed against S. Typhi-inoculated stool samples. mPCR was able to detect S. Typhi cell suspension of 1x105 cfu/mL. We found that modified multiplex PCR was useful to detect S. Typhi from stool samples within 24h whereas it takes 3-5days to detect by classic diagnosis method.

Keyword

Salmonella Typhi; Multiplex PCR; invA; fliC-d; viaB

MeSH Terms

Citrobacter freundii
Communicable Diseases
Developing Countries
Multiplex Polymerase Chain Reaction
Public Health
Salmonella
Salmonella enterica
Salmonella typhi
Sensitivity and Specificity
Typhoid Fever

Figure

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  Figure 1 The specificity and sensitivity of modified multiplex PCR to detect S. Typhi. (A) Specificity of multiplex PCR. invA gene was amplified all Salmonella serovars but viaB and fliC-d was only S. Typhi by modified multiplex PCR. All Salmonella serovar was inoculated into human feces, concentration of 108 cfu/mL. M. 100 bp DNA ladder; Lane 1. S. Enteritidis; Lane 2. S. Typhimurium; Lane 3. S. Typhi; Lane 4, S. Braenderup; Lane 5, S. London; Lane 6. S. ParatyphiA; Lane 7, S. Infantis; Lane 8, S. Montevideo; Lane 9, Negative control (distiled water). (B) Sensitivity of multiplex PCR. Modified multiplex PCR was able to detect 105 cfu/mL of S. Typhi, which contained human feces. M, 100bp DNA ladder; Lane 1, 107 cfu/mL; Lane 2, 106 cfu/mL; Lane 3, 105 cfu/mL; Lane 4, 104 cfu/mL; Lane 5, 103 cfu/mL; Lane 6, 102 cfu/mL; Lane 7, 101 cfu/mL; Lane 8, 100 cfu/mL; Lane 9, 0 cfu/mL; Lane 10, Positive control; Lane 11, Negative control (distiled water).

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