Abstract
Enterotoxigenic Escherichia coli (ETEC) expressing the colonization factor CS6 is widespread in many developing countries, including India. The different allelic variants of CS6, caused by point mutations in its structural genes, cssA and cssB, are designated AIBI, AIIBII, AIIIBI, AIBII, and AIIIBII. A simple, reliable, and specific mismatch amplificationmutation assay based on real-time quantitative PCR (MAMA-qPCR) was developed for the first time for the detection of CS6-expressing ETEC, along with the identification of allelic variations. The assay was based on mismatched nucleotide incorporation at the penultimate base at the 3′ ends of the reverse primers specific for cssA and cssB and was validated using 38 CS6-expressing ETEC isolates. This strategy was effective in detecting all the alleles containing single-nucleotide polymorphisms. Using MAMA-qPCR, we also tested CS6 allelic variants in 145 ETEC isolates from children with acute diarrhea and asymptomatic infections, with the latter serving as controls. We observed that the AIBI and AIIIBI allelic variants were mostly associated with cases rather than controls, whereas the AIIBII variants were detected mostly in controls. In addition, the AIBI and AIIIBI alleles were frequently associated with ETEC harboring the heat-stable toxin gene (est) alone or with the heat-labile toxin gene (elt), whereas the AIIBII allele was predominant in ETEC isolates harboring the elt gene. This study may help in understanding the association of allelic variants in CS6-expressing ETEC with the clinical features of diarrhea, as well as in ETEC vaccine studies.
| Original language | English |
|---|---|
| Pages (from-to) | 1308-1312 |
| Number of pages | 5 |
| Journal | Journal of Clinical Microbiology |
| Volume | 50 |
| Issue number | 4 |
| DOIs | |
| Publication status | Published - Apr 2012 |
| Externally published | Yes |
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ASJC Scopus subject areas
- Microbiology (medical)
Cite this
Real-time PCR-based mismatch amplification mutation assay for specific detection of CS6-expressing allelic variants of enterotoxigenic Escherichia coli and its application in assessing diarrheal cases and asymptomatic controls. / Sabui, Subrata; Dutta, Sanjucta; Debnath, Anusuya; Ghosh, Avishek; Hamabata, T.; Rajendran, K.; Ramamurthy, T.; Nataro, James P.; Sur, Dipika; Levine, Myron M.; Chatterjee, Nabendu Sekhar.
In: Journal of Clinical Microbiology, Vol. 50, No. 4, 04.2012, p. 1308-1312.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Real-time PCR-based mismatch amplification mutation assay for specific detection of CS6-expressing allelic variants of enterotoxigenic Escherichia coli and its application in assessing diarrheal cases and asymptomatic controls
AU - Sabui, Subrata
AU - Dutta, Sanjucta
AU - Debnath, Anusuya
AU - Ghosh, Avishek
AU - Hamabata, T.
AU - Rajendran, K.
AU - Ramamurthy, T.
AU - Nataro, James P.
AU - Sur, Dipika
AU - Levine, Myron M.
AU - Chatterjee, Nabendu Sekhar
PY - 2012/4
Y1 - 2012/4
N2 - Enterotoxigenic Escherichia coli (ETEC) expressing the colonization factor CS6 is widespread in many developing countries, including India. The different allelic variants of CS6, caused by point mutations in its structural genes, cssA and cssB, are designated AIBI, AIIBII, AIIIBI, AIBII, and AIIIBII. A simple, reliable, and specific mismatch amplificationmutation assay based on real-time quantitative PCR (MAMA-qPCR) was developed for the first time for the detection of CS6-expressing ETEC, along with the identification of allelic variations. The assay was based on mismatched nucleotide incorporation at the penultimate base at the 3′ ends of the reverse primers specific for cssA and cssB and was validated using 38 CS6-expressing ETEC isolates. This strategy was effective in detecting all the alleles containing single-nucleotide polymorphisms. Using MAMA-qPCR, we also tested CS6 allelic variants in 145 ETEC isolates from children with acute diarrhea and asymptomatic infections, with the latter serving as controls. We observed that the AIBI and AIIIBI allelic variants were mostly associated with cases rather than controls, whereas the AIIBII variants were detected mostly in controls. In addition, the AIBI and AIIIBI alleles were frequently associated with ETEC harboring the heat-stable toxin gene (est) alone or with the heat-labile toxin gene (elt), whereas the AIIBII allele was predominant in ETEC isolates harboring the elt gene. This study may help in understanding the association of allelic variants in CS6-expressing ETEC with the clinical features of diarrhea, as well as in ETEC vaccine studies.
AB - Enterotoxigenic Escherichia coli (ETEC) expressing the colonization factor CS6 is widespread in many developing countries, including India. The different allelic variants of CS6, caused by point mutations in its structural genes, cssA and cssB, are designated AIBI, AIIBII, AIIIBI, AIBII, and AIIIBII. A simple, reliable, and specific mismatch amplificationmutation assay based on real-time quantitative PCR (MAMA-qPCR) was developed for the first time for the detection of CS6-expressing ETEC, along with the identification of allelic variations. The assay was based on mismatched nucleotide incorporation at the penultimate base at the 3′ ends of the reverse primers specific for cssA and cssB and was validated using 38 CS6-expressing ETEC isolates. This strategy was effective in detecting all the alleles containing single-nucleotide polymorphisms. Using MAMA-qPCR, we also tested CS6 allelic variants in 145 ETEC isolates from children with acute diarrhea and asymptomatic infections, with the latter serving as controls. We observed that the AIBI and AIIIBI allelic variants were mostly associated with cases rather than controls, whereas the AIIBII variants were detected mostly in controls. In addition, the AIBI and AIIIBI alleles were frequently associated with ETEC harboring the heat-stable toxin gene (est) alone or with the heat-labile toxin gene (elt), whereas the AIIBII allele was predominant in ETEC isolates harboring the elt gene. This study may help in understanding the association of allelic variants in CS6-expressing ETEC with the clinical features of diarrhea, as well as in ETEC vaccine studies.
UR - http://www.scopus.com/inward/record.url?scp=84858612625&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=84858612625&partnerID=8YFLogxK
U2 - 10.1128/JCM.05424-11
DO - 10.1128/JCM.05424-11
M3 - Article
C2 - 22219305
AN - SCOPUS:84858612625
VL - 50
SP - 1308
EP - 1312
JO - Journal of Clinical Microbiology
JF - Journal of Clinical Microbiology
SN - 0095-1137
IS - 4
ER -