Comparative Evaluation of an Automated Repetitive-Sequence-Based PCR Instrument versus Pulsed-Field Gel Electrophoresis in the Setting of a Serratia marcescens Nosocomial Infection Outbreak

Abstract

A semiautomated, repetitive-sequence-based PCR (rep-PCR) instrument (DiversiLab system) was evaluated in comparison with pulsed-field gel electrophoresis (PFGE) to investigate an outbreak of Serratia marcescens infections in a neonatal intensive care unit (NICU). A selection of 36 epidemiologically related and 8 epidemiologically unrelated isolates was analyzed. Among the epidemiologically related isolates, PFGE identified five genetically unrelated patterns. Thirty-two isolates from patients and wet nurses showed the same PFGE profile (pattern A). Genetically unrelated PFGE patterns were found in one patient (pattern B), in two wet nurses (patterns C and D), and in an environmental isolate from the NICU (pattern G). Rep-PCR identified seven different patterns, three of which included the 32 isolates of PFGE type A. One or two band differences in isolates of these three types allowed isolates to be categorized as similar and included in a unique cluster. Isolates of different PFGE types were also of unrelated rep-PCR types. All of the epidemiologically unrelated isolates were of different PFGE and rep-PCR types. The level of discrimination exhibited by rep-PCR with the DiversiLab system allowed us to conclude that this method was able to identify genetic similarity in a spatio-temporal cluster of S. marcescens isolates.Concerns regarding the emergence of hospital-acquired infections, increasing antimicrobial resistance, and the increase in morbidity, mortality, and costs associated with these infections drive the need for refinement of molecular approaches to aid in the diagnosis and epidemiological analysis of nosocomial infections.Several methods based on DNA analysis are now available that provide information about the genetic relatedness of isolates of the same species (26). These DNA-based molecular methodologies include pulsed-field gel electrophoresis (PFGE), PCR-based typing methods, and multilocus sequence analysis.

Phenotypic and Molecular Characteristics of Carbapenem-Non-Susceptible Enterobacteriaceae from a Teaching Hospital in Wenzhou, Southern China

“…The genomic DNA of these isolates bearing the carbapenem-resistance genes was analyzed by PFGE. K. pneumoniae genomic DNA was digested with XbaI (24) while that of S. marcescens was digested with SpeI (25). The DNA fragments were separated using a CHEF-Mapper XA PFGE system (Bio-Rad, Hercules, Calif., USA) for 22 h at 6 V/cm and 149 C, with a pulse angle of 1209and pulse duration from 5 s to 25 s. The restriction patterns were analyzed and interpreted according to the criteria proposed by Tenover et al (26).…”

Controlled Performance Evaluation of the DiversiLab Repetitive-Sequence-Based Genotyping System for Typing Multidrug-Resistant Health Care-Associated Bacterial Pathogens

“…The DL system is a rapid and semiautomated rep-PCR commercial system that uses standard PCR kits and high-resolution amplicon detection by microfluidic capillary electrophoresis. Its discriminatory power and concordance with other typing methods have been previously evaluated for a number of bacterial pathogens (5,7,11,13,16,19,21,23,24). A recent study analyzed its usefulness in identifying hospital outbreaks caused by different bacterial pathogens (12).…”

Applicability of DiversiLab Repetitive Sequence-Based PCR Method in Epidemiological Typing of EnterohemorrhagicEscherichia coli(EHEC)

“…Recently, a commercial system for fingerprinting strains by the rep-PCR method (DiversiLab, Bacterial Barcodes, Athens, GA) has become available, offering both standardization and automated data analysis. This approach has been successfully used with several bacterial genera/species, such as Salmonella, Serratia, Pseudomonas aeruginosa, methicillin-resistant Staphylococcus aureus, Enterococcus, and Clostridium difficile (Ross et al, 2005;Doléans-Jordheim et al, 2009;Ben-Darif et al, 2010;Chuang et al, 2010;Fluit et al, 2010;Grisold et al, 2010;Ligozzi et al, 2010;Pasanen et al, 2011). In addition, several E. coli strains such as uropathogenic E. coli, extended-spectrum b-lactamase, and AmpC-producing E. coli and strains isolated from neonatal meningitis cases have been analyzed with the DiversiLab method (Bonacorsi et al, 2009;Pitout et al, 2009;Bogaerts et al, 2010;Brolund et al, 2010;Lau et al, 2010).…”