Evaluation of restriction endonuclease fingerprinting of chromosomal DNA and plasmid profile analysis for characterization of multiresistant coagulase-negative staphylococci in bacteremic neonates

Abstract

A procedure was developed for restriction endonuclease fingerprinting (REF) of the chromosomal DNA of coagulase-negative staphylococci. A total of 48 isolates comprising 29 Staphylococcus epidermidis and 19 Staphylococcus haemolyticus isolates from blood and mucocutaneous sites of 15 premature neonates were characterized by REF, plasmid profile (PP) analysis, antimicrobial susceptibility testing, biotyping, and slime production. On the basis of REF analysis of chromosomal DNA, the 48 coagulase-negative staphylococcal isolates were subdivided into 10 subgroups, whereas PP analysis subdivided the strains into 20 distinct subgroups. REF analysis of total DNA (i.e., chromosome plus plasmid) resulted in the same 20 subgroups as were subdivided by PP analysis. The high discriminatory power of PP analysis was associated with the variability of plasmid content in coagulase-negative staphylococcal strains isolated during the outbreak. REF patterns were found to be stable both in vitro and in vivo. Isolates carried from 2 to 10 plasmids that ranged in molecular size from 0.9 to 39.5 megadaltons. Plasmids were disseminated among the coagulase-negative staphylococci, regardless of the genetic relatedness of their chromosomal DNAs. Hence, a lack of correlation existed between the grouping of isolates by REF analysis of chromosomal DNA and the grouping by PP analysis. There were one and two distinct chromosomal patterns among 4 of 4 blood cultures and 15 of 15 mucocutaneous cultures of S. haemolyticus, respectively. In contrast, a higher proportion of distinct chromosomal patterns was found for S. epidermidis in blood cultures (7 of 11 cultures) compared with those identified for isolates in mucocutaneous cultures (6 of 18 cultures). In summary, REF analysis of chromosomal DNA, rather than total DNA, is a useful marker for epidemiological investigations of coagulase-negative staphylococci. PP analysis can also be used to provide additional epidemiological information regarding the most recent genetic events.