A Pentaplex PCR Assay for the Detection and Differentiation of Shigella Species
The magnitude of shigellosis in developing countries is largely unknown because an affordable detection method is not available. Current laboratory diagnosis of Shigella spp. is laborious and time consuming and has low sensitivity. Hence, in the present study, a molecular-based diagnostic assay wh...
| Main Authors: | , , , |
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| Format: | Article |
| Language: | English |
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Hindawi Publishing Corporation
2013
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| Online Access: | http://eprints.usm.my/38094/1/A_Pentaplex_PCR_Assay_for_the_Detection_and_Differentiation_of_Shigella_Species.pdf |
| _version_ | 1825833785857409024 |
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| author | Ojha, Suvash Chandra Chan, Yean Yean Ismail, Asma Banga Singh, Kirnpal Kaur |
| author_facet | Ojha, Suvash Chandra Chan, Yean Yean Ismail, Asma Banga Singh, Kirnpal Kaur |
| author_sort | Ojha, Suvash Chandra |
| collection | USM |
| description | The magnitude of shigellosis in developing countries is largely unknown because an affordable detection method is not available.
Current laboratory diagnosis of Shigella spp. is laborious and time consuming and has low sensitivity. Hence, in the present study,
a molecular-based diagnostic assay which amplifies simultaneously four specific genes to identify invC for Shigella genus, rfc for S.
flexneri, wbgZ for S. sonnei, andrfpB for S. dysenteriae, aswell as one internal control (ompA) gene, was developed in a single reaction
to detect and differentiate Shigella spp. Validation with 120 Shigella strains and 37 non-Shigella strains yielded 100% specificity. The
sensitivity of the PCR was 100 pg of genomic DNA, 5.4 × 104 CFU/ml, or approximately 120 CFU per reaction mixture of bacteria.
The sensitivity of the pentaplex PCR assay was further improved following preincubation of the stool samples in Gram-negative
broth. A preliminary study with 30 diarrhoeal specimens resulted in no cross-reaction with other non-Shigella strains tested. We
conclude that the developed pentaplex PCR assay is robust and can provide information about the four target genes that are essential
for the identification of the Shigella genus and the three Shigella species responsible for the majority of shigellosis cases. |
| first_indexed | 2024-03-06T15:13:00Z |
| format | Article |
| id | usm.eprints-38094 |
| institution | Universiti Sains Malaysia |
| language | English |
| last_indexed | 2024-03-06T15:13:00Z |
| publishDate | 2013 |
| publisher | Hindawi Publishing Corporation |
| record_format | dspace |
| spelling | usm.eprints-380942017-12-22T08:07:27Z http://eprints.usm.my/38094/ A Pentaplex PCR Assay for the Detection and Differentiation of Shigella Species Ojha, Suvash Chandra Chan, Yean Yean Ismail, Asma Banga Singh, Kirnpal Kaur QR1-502 Microbiology R735-854 Medical education. Medical schools. Research The magnitude of shigellosis in developing countries is largely unknown because an affordable detection method is not available. Current laboratory diagnosis of Shigella spp. is laborious and time consuming and has low sensitivity. Hence, in the present study, a molecular-based diagnostic assay which amplifies simultaneously four specific genes to identify invC for Shigella genus, rfc for S. flexneri, wbgZ for S. sonnei, andrfpB for S. dysenteriae, aswell as one internal control (ompA) gene, was developed in a single reaction to detect and differentiate Shigella spp. Validation with 120 Shigella strains and 37 non-Shigella strains yielded 100% specificity. The sensitivity of the PCR was 100 pg of genomic DNA, 5.4 × 104 CFU/ml, or approximately 120 CFU per reaction mixture of bacteria. The sensitivity of the pentaplex PCR assay was further improved following preincubation of the stool samples in Gram-negative broth. A preliminary study with 30 diarrhoeal specimens resulted in no cross-reaction with other non-Shigella strains tested. We conclude that the developed pentaplex PCR assay is robust and can provide information about the four target genes that are essential for the identification of the Shigella genus and the three Shigella species responsible for the majority of shigellosis cases. Hindawi Publishing Corporation 2013 Article PeerReviewed application/pdf en http://eprints.usm.my/38094/1/A_Pentaplex_PCR_Assay_for_the_Detection_and_Differentiation_of_Shigella_Species.pdf Ojha, Suvash Chandra and Chan, Yean Yean and Ismail, Asma and Banga Singh, Kirnpal Kaur (2013) A Pentaplex PCR Assay for the Detection and Differentiation of Shigella Species. BioMed Research International, 2013 (412370). pp. 1-9. ISSN 2314-6133 http://dx.doi.org/10.1155/2013/412370 |
| spellingShingle | QR1-502 Microbiology R735-854 Medical education. Medical schools. Research Ojha, Suvash Chandra Chan, Yean Yean Ismail, Asma Banga Singh, Kirnpal Kaur A Pentaplex PCR Assay for the Detection and Differentiation of Shigella Species |
| title | A Pentaplex PCR Assay for the Detection and Differentiation of Shigella Species |
| title_full | A Pentaplex PCR Assay for the Detection and Differentiation of Shigella Species |
| title_fullStr | A Pentaplex PCR Assay for the Detection and Differentiation of Shigella Species |
| title_full_unstemmed | A Pentaplex PCR Assay for the Detection and Differentiation of Shigella Species |
| title_short | A Pentaplex PCR Assay for the Detection and Differentiation of Shigella Species |
| title_sort | pentaplex pcr assay for the detection and differentiation of shigella species |
| topic | QR1-502 Microbiology R735-854 Medical education. Medical schools. Research |
| url | http://eprints.usm.my/38094/1/A_Pentaplex_PCR_Assay_for_the_Detection_and_Differentiation_of_Shigella_Species.pdf |
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