Abstract
For specific detection of the probiotic Bifidobacterium sp. strain LW420 in infant feces and for rapid quality control of this strain in culture, three strain-specific 16S rRNA gene-targeted primers have been developed. These primers allow specific detection of the organism via PCR. Specificity of the primers was determined in DNA samples isolated from single-strain and mixed cultures of bifidobacteria and in heterogenous fecal samples. The feasibility of this method for use in specific detection of probiotic strains was investigated through addition of Bifidobacterium sp. strain LW420 to infant instant milk formula (IMF) and PCR analyses of bacterial DNA isolated from feces of 17 newborn IMF-fed infants. In feces of all nine babies that had been fed with the probiotic IMF, the strain-specific PCR signal could be detected. No signal was found in feces of the eight infants that had been fed with a nonprobiotic IMF, demonstrating the specificity of the PCR method. All 17 infants developed a major fecal Bifidobacterium population already after 3 days, as determined through genus-specific and strain-specific PCR. Phenotypical screening of Bifidobacterium sp. strain LW420 and analysis of homology of the 16S rRNA gene sequence of this strain with that of other bifidobacteria deposited in databases do not allow positive classification of LW420 among the currently known species of Bifidobacterium.
MeSH Terms
Animals
Bifidobacterium/genetics,growth & development,isolation & purification
DNA Primers
DNA, Ribosomal/genetics
Double-Blind Method
Feces/microbiology
Fermentation
Humans
Infant Food/microbiology
Infant, Newborn
Milk
Molecular Sequence Data
Polymerase Chain Reaction/methods
RNA, Ribosomal, 16S/genetics
Sensitivity and Specificity
Sequence Analysis, DNA
Sequence Homology, Nucleic Acid
Chemicals
DNA Primers
DNA, Ribosomal
RNA, Ribosomal, 16S
Authors & Affiliations
6 authors, click to expand affiliations / ORCID
Kok R G
Department of Microbiology, E. C. Slater Institute, BioCentrum Amsterdam, The Netherlands.
de Waal A
Schut F
Welling G W
Weenk G
Hellingwerf K J
References (15)
15 references, click to expand
-
The effect of "humanised" milks and supplemented breast feeding on the faecal flora of infants.
J Med Microbiol. 1977 Nov;10(4):403-13
PMID: 21296
-
DNA sequencing with chain-terminating inhibitors.
Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463-7
PMID: 271968
-
Bacterial evolution.
Microbiol Rev. 1987 Jun;51(2):221-71
PMID: 2439888
-
Improved tools for biological sequence comparison.
Proc Natl Acad Sci U S A. 1988 Apr;85(8):2444-8
PMID: 3162770
-
Rapid and simple method for purification of nucleic acids.
J Clin Microbiol. 1990 Mar;28(3):495-503
PMID: 1691208
-
The Ribosomal Database Project.
Nucleic Acids Res. 1992 May 11;20 Suppl:2199-200
PMID: 1598241
-
Rapid identification of bacteria on the basis of polymerase chain reaction-amplified ribosomal DNA spacer polymorphisms.
Appl Environ Microbiol. 1993 Apr;59(4):945-52
PMID: 8476298
-
Sequence-based differentiation of strains in the Mycobacterium avium complex.
J Bacteriol. 1993 May;175(10):2818-25
PMID: 8491701
-
Phylogenetic identification and in situ detection of individual microbial cells without cultivation.
Microbiol Rev. 1995 Mar;59(1):143-69
PMID: 7535888
-
Design and evaluation of Bacteroides DNA probes for the specific detection of human fecal pollution.
Appl Environ Microbiol. 1995 Apr;61(4):1171-9
PMID: 7538270
-
Isolation of human immunodeficiency virus type 1 (HIV-1) RNA from feces by a simple method and difference between HIV-1 subpopulations in feces and serum.
J Clin Microbiol. 1995 Mar;33(3):581-8
PMID: 7751361
-
Quantitative fluorescence in situ hybridization of Bifidobacterium spp. with genus-specific 16S rRNA-targeted probes and its application in fecal samples.
Appl Environ Microbiol. 1995 Aug;61(8):3069-75
PMID: 7487040
-
16S rRNA and 16S to 23S internal transcribed spacer sequence analyses reveal inter- and intraspecific Bifidobacterium phylogeny.
Int J Syst Bacteriol. 1996 Jan;46(1):102-11
PMID: 8573484
-
Phylogenetic analysis of Fusobacterium prausnitzii based upon the 16S rRNA gene sequence and PCR confirmation.
Int J Syst Bacteriol. 1996 Jan;46(1):341-3
PMID: 8573517
-
Microbiological assay for thiamin using Lactobacillus viridescens.
J Bacteriol. 1957 Dec;74(6):818-21
PMID: 13502311