Quality control for bacterial inhibition assays: DNA fingerprinting of microorganisms by rep-PCR

Abstract

The bacterial inhibition assay (BIA) has been useful in a variety of newborn screening tests. BIAs rely on specific bacterial indicator strains as standardized components in different assays. Recently, DNA fingerprinting methods have been developed for eubacteria which utilize conserved repetitive DNA sequences. The repetitive DNA sequences, REP (Repetitive Extragenic Palindromic) and ERIC (Enterobacterial Repetitive Intergenic Consensus), can be used as oligonucleotide primer binding sites for the polymerase chain reaction (PCR) amplification of unique DNA sequences located between the repeats. Repetitive sequence based PCR, generally known as rep-PCR, allows the production of species- and strain-specific genomic fingerprint patterns following size-fractionation of the amplified DNA fragments by agarose gel electrophoresis. Two Bacillus subtilis strains, ATCC 6051 and 6633, currently used in newborn screening BlAs were distinguished by rep-PCR. Using rep-PCR to fingerprint bacterial strains collected from different newborn screening laboratories, an anomalous sample was revealed which yielded rep-PCR fingerprint patterns distinct from other laboratory samples of the same strain. Genomic fingerprinting by rep-PCR may be useful for quality control of specific bacterial strains used in BIAs.