Molecular typing of Klebsiella pneumoniae in neonatal intensive care units using amplified fragment length polymorphism analysis

Abstract number: R2158

Denks K., Parm Ü., Huik K., Avi R., Metsvaht T., Karki T., Lutsar I.

Objectives: The aim of the study was to determine the population dynamics of Klebsiella pneumoniae in neonatal intensive care units (NICUs) of two major hospitals in Estonia, using amplified fragment length polymorphism (AFLP) analysis.

Methods: From August 2006 until December 2007, repeated perirectal and nasopharyngeal swabs were collected from all neonates admitted to two Estonian NICUs with suspected early onset neonatal sepsis. Mucosal carriage of K. pneumoniae was detected in 51 of 278 eligible patients. Finally, 47 of 51 patients were analysed: 38 in unit A (including 5 patients with blood stream infection) and 9 in unit B (plus 1 patient with blood stream infection but without mucosal colonisation). Depending on the length of hospitalisation, the per patient isolate number varied from 1 to 7. Alltogether, 88 perirectal, 52 nasopharyngal, 3 tracheal and 5 blood isolates were used. The AFLP analysis was implemented as described previously1. After restriction enzyme digestion, the fragments were selectively amplified and separated in agarose gel electrophoresis. The fingerprints on gel were analysed using GeneTools program (Syngene). In parallel, 10 samples of 5 patients were typed using pulse-field gel electrophoresis (PFGE) technique.

Results: After the reproducibility analysis of AFLP method, isolates with up to 3-band pattern difference were considered identical. Two predominant K. pneumoniae clonal groups (type A in 26 patients over 7 months and five months later, type K in 6 patients over 2 months) were detected in unit A, while in unit B all colonising K. pneumoniae strains were different. In all but one subject within individual concordance between nasopharyngeal and perirectal isolates was observed. All five blood stream infections in unit A occurred during the period of type A K. pneumoniae colonisation. On AFLP and PFGE analysis, all five invasive isolates were identical to colonising ones. Ampicillin and cefotaxime resistance was detected in 41% and 1.4% of isolates, respectively, but was not reflected on AFLP fingerprints.

Conclusion: AFLP method identified two predominant colonising clonal groups of K. pneumoniae in NICU setting A, whereas in setting B all patients had different strain. Vast majority of invasive disease was caused by only type A K. pneumoniae infection, but it is not known whether because of possibly higher virulence or as a result of widespread colonisation.

References

1. Van der Zee et al 2003. J. Clin. Microbiol. 41:798–802