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Original Article
  |     |   Neonatal Med_20_2_189_198.pdf
Neonatal Med May;20(2):189-198.
Published online 2013 June 26   doi: https://doi.org/10.5385/nm.2013.20.2.189
Copyright ⓒ 2013 Neonatal Medicine Neonatal Medicine
Denaturing Gradient Gel Electrophoresis for Detection of Bacterial Population and Diversity in Amniotic Fluids and Neonatal Gastric Fluids
Young Don Kim, M.D., Sun Nyoung Yu*, Seong Chol Kim, M.D.†, and Soon Cheol Ahn, Ph.D.*
Department of Pediatrics, Jeju National University School of Medicine, Jeju, Korea
Department of Microbiology and Immunology*, Pusan National University School of Medicine, Pusan, Korea
Department of Obstetrics and Gynecology†, Pusan National University School of Medicine, Pusan, Korea
Corresponding Author: Soon Cheol Ahn, Ph.D. , Tel: +82-51-510-8099 , Fax: +82-55-382-8090 , Email: ahnsc@pusan.ac.kr
ABSTRACT
Purpose: Intra-amniotic infection (IAI) is often polymicrobial, and the 16S rDNA PCR assay has a major limitation that its interpretation is difficult in the presence of multiple 16S rDNAs. Denaturing gradient gel electrophoresis (DGGE) can overcome this limitation by separating PCR products based on sequence. We performed the DGGE analysis to investigate bacterial preva-lence and diversity in amniotic fluids from pregnant women with preterm births and gastric fluids from their newborns.
Methods: DNA was extracted from bacterial cells in amniotic fluid (AF) and gastric fluid (GF) and was amplified with universal 16S rDNA primers. For DGGE analysis, the PCR products were loaded onto polyacrylamide gels that were made with denaturing gradients.
Results: Bacterial 16S rDNA was detected by PCR from all AF and GF samples. The bacterial species in AF samples were the following: Lactobacillus reuteri (87.0%), uncultured Enterococcus species (65.2%), Ureaplasma urealyticum (13.0%), and Enterococcus faecalis (4.3%). The bacterial species in GF samples were the following: Lactobacillus reuteri (95.2%), uncultured Enterococcus species (42.9%), and Ureaplasma urealyticum (4.8%). Two or more species were identified from 69.6% of AF and 47.6% of GF samples.
Conclusion: We suggest that DGGE analysis allows improved understanding of microbial diversity and community in AF and GF.
Keywords: Preterm infant, Amniotic fluid, Gastric fluid, Intra-amniotic, Infection, Denaturing gradient gel electrophoresis, Bacterial diversity
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