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Narciss Okhravi, Peter Adamson, Melville M. Matheson, Hamish M. A. Towler, Susan Lightman; PCR-RFLP–Mediated Detection and Speciation of Bacterial Species Causing Endophthalmitis. Invest. Ophthalmol. Vis. Sci. 2000;41(6):1438-1447.
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purpose. To determine the usefulness of polymerase chain reaction–restriction
fragment length polymorphism (PCR-RFLP) in the identification and
speciation of bacteria causing endophthalmitis.
methods. PCR-RFLP was performed on 53 strains of 14 bacterial species (eight
Gram positive and five Gram negative) collected from both keratitis and
endophthalmitis patients. Two pairs of oligonucleotide primers based on
the 16S rDNA gene were used to PCR-amplify 1.2- and 1.0-kb fragments of
bacterial genomic DNA. RFLPs within the PCR product were used to
speciate the organisms.
results. The sensitivity of the nested PCR amplification reaction was one
organism. All bacteria tested could be identified and speciated using
RFLP analysis except for Escherichia coli and Serratia marcescens, which could not be
interdifferentiated using RFLP. Molecular analysis of two vitreous
samples from two eyes with typical signs of bacterial endophthalmitis
confirmed the presence of E. coli in the vitreous from a
culture-positive case with E. coli endophthalmitis and
revealed the presence of Staphylococcus epidermidis in
the vitreous of a culture-negative case.
conclusions. It is expected that this technique will provide a useful laboratory
tool for future microbiologic diagnosis of patients presenting with
endophthalmitis, especially for those eyes that prove culture
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