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J Clin Microbiol. May 1996; 34(5): 1193–1202.
PMCID: PMC228981

Validation of use of whole-cell repetitive extragenic palindromic sequence-based PCR (REP-PCR) for typing strains belonging to the Acinetobacter calcoaceticus-Acinetobacter baumannii complex and application of the method to the investigation of a hospital outbreak.


Acinetobacter spp. are being reported with increasing frequency as causes of nosocomial infection. In order to identify reservoirs of infection as quickly as possible, a rapid typing method that can differentiate epidemic strains from environmental and nonepidemic strains is needed. In 1993, a cluster of Acinetobacter baumannii isolates from five patients in the adult intensive therapy unit of our tertiary-care teaching hospital led us to develop and optimize a rapid repetitive extragenic palindromic sequence-based PCR (REP-PCR) typing protocol for members of the Acinetobacter calcoaceticus-A. baumannii complex that uses boiled colonies and consensus primers aimed at repetitive extragenic palindromic sequences. Four of the five patient isolates gave the same REP-PCR typing pattern as isolates of A. baumannii obtained from the temperature probe of a Bennett humidifier; the fifth isolate had a unique profile. Disinfection of the probe with 70% ethanol, as recommended by the manufacturer, proved ineffective, as A. baumannii with the same REP-PCR pattern was isolated from it 10 days after cleaning, necessitating a change in our decontamination procedure. Results obtained with REP-PCR were subsequently confirmed by ribotyping. To evaluate the discriminatory power (D) of REP-PCR for typing members of the A. calcoaceticus-A. baumannii complex, compared with that of ribotyping, we have applied both methods to a collection of 85 strains that included representatives of six DNA groups within the complex. Ribotyping using EcoRI digests yielded 53 patterns (D = 0.98), whereas 68 different REP-PCR patterns were observed (D = 0.99). By computer-assisted analysis of gel images, 74 patterns were observed with REP-PCR (D = 1.0). Overall, REP-PCR typing proved to be slightly more discriminatory than ribotyping. Our results indicate that REP-PCR typing used boiled colonies is a simple, rapid, and effective means of typing members of the A. calcoaceticus-A. baumannii complex.

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Selected References

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  • Al-Khoja MS, Darrell JH. The skin as the source of Acinetobacter and Moraxella species occurring in blood cultures. J Clin Pathol. 1979 May;32(5):497–499. [PMC free article] [PubMed]
  • Beck-Sagué CM, Jarvis WR, Brook JH, Culver DH, Potts A, Gay E, Shotts BW, Hill B, Anderson RL, Weinstein MP. Epidemic bacteremia due to Acinetobacter baumannii in five intensive care units. Am J Epidemiol. 1990 Oct;132(4):723–733. [PubMed]
  • Bell DA, DeMarini DM. Excessive cycling converts PCR products to random-length higher molecular weight fragments. Nucleic Acids Res. 1991 Sep 25;19(18):5079–5079. [PMC free article] [PubMed]
  • Bergogne-Berezin E, Joly-Guillou ML. An underestimated nosocomial pathogen, Acinetobacter calcoaceticus. J Antimicrob Chemother. 1985 Nov;16(5):535–538. [PubMed]
  • Bergogne-Bérézin E, Joly-Guillou ML, Vieu JF. Epidemiology of nosocomial infections due to Acinetobacter calcoaceticus. J Hosp Infect. 1987 Sep;10(2):105–113. [PubMed]
  • Berthelot P, Grattard F, Mahul P, Jospe R, Pozzetto B, Ros A, Gaudin OG, Auboyer C. Ventilator temperature sensors: an unusual source of Pseudomonas cepacia in nosocomial infection. J Hosp Infect. 1993 Sep;25(1):33–43. [PubMed]
  • Bouvet PJ, Jeanjean S. Delineation of new proteolytic genomic species in the genus Acinetobacter. Res Microbiol. 1989 May-Jun;140(4-5):291–299. [PubMed]
  • Cefai C, Richards J, Gould FK, McPeake P. An outbreak of Acinetobacter respiratory tract infection resulting from incomplete disinfection of ventilatory equipment. J Hosp Infect. 1990 Feb;15(2):177–182. [PubMed]
  • Contant J, Kemeny E, Oxley C, Perry E, Garber G. Investigation of an outbreak of Acinetobacter calcoaceticus var. anitratus infections in an adult intensive care unit. Am J Infect Control. 1990 Aug;18(4):288–291. [PubMed]
  • de Bruijn FJ. Use of repetitive (repetitive extragenic palindromic and enterobacterial repetitive intergeneric consensus) sequences and the polymerase chain reaction to fingerprint the genomes of Rhizobium meliloti isolates and other soil bacteria. Appl Environ Microbiol. 1992 Jul;58(7):2180–2187. [PMC free article] [PubMed]
  • Dijkshoorn L, Aucken HM, Gerner-Smidt P, Kaufmann ME, Ursing J, Pitt TL. Correlation of typing methods for Acinetobacter isolates from hospital outbreaks. J Clin Microbiol. 1993 Mar;31(3):702–705. [PMC free article] [PubMed]
  • Fagon JY, Chastre J, Domart Y, Trouillet JL, Pierre J, Darne C, Gibert C. Nosocomial pneumonia in patients receiving continuous mechanical ventilation. Prospective analysis of 52 episodes with use of a protected specimen brush and quantitative culture techniques. Am Rev Respir Dis. 1989 Apr;139(4):877–884. [PubMed]
  • Gerner-Smidt P. Endemic occurrence of Acinetobacter calcoaceticus biovar anitratus in an intensive care unit. J Hosp Infect. 1987 Nov;10(3):265–272. [PubMed]
  • Gerner-Smidt P. Frequency of plasmids in strains of Acinetobacter calcoaceticus. J Hosp Infect. 1989 Jul;14(1):23–28. [PubMed]
  • Gerner-Smidt P. Ribotyping of the Acinetobacter calcoaceticus-Acinetobacter baumannii complex. J Clin Microbiol. 1992 Oct;30(10):2680–2685. [PMC free article] [PubMed]
  • Gerner-Smidt P, Tjernberg I. Acinetobacter in Denmark: II. Molecular studies of the Acinetobacter calcoaceticus-Acinetobacter baumannii complex. APMIS. 1993 Nov;101(11):826–832. [PubMed]
  • Gerner-Smidt P, Tjernberg I, Ursing J. Reliability of phenotypic tests for identification of Acinetobacter species. J Clin Microbiol. 1991 Feb;29(2):277–282. [PMC free article] [PubMed]
  • Gervich DH, Grout CS. An outbreak of nosocomial Acinetobacter infections from humidifiers. Am J Infect Control. 1985 Oct;13(5):210–215. [PubMed]
  • Getchell-White SI, Donowitz LG, Gröschel DH. The inanimate environment of an intensive care unit as a potential source of nosocomial bacteria: evidence for long survival of Acinetobacter calcoaceticus. Infect Control Hosp Epidemiol. 1989 Sep;10(9):402–407. [PubMed]
  • Goodhart GL, Abrutyn E, Watson R, Root RK, Egert J. Community-acquired Acinetobacter calcoaceticus var anitratus pneumonia. JAMA. 1977 Oct 3;238(14):1516–1518. [PubMed]
  • Gouby A, Carles-Nurit MJ, Bouziges N, Bourg G, Mesnard R, Bouvet PJ. Use of pulsed-field gel electrophoresis for investigation of hospital outbreaks of Acinetobacter baumannii. J Clin Microbiol. 1992 Jun;30(6):1588–1591. [PMC free article] [PubMed]
  • Gräser Y, Klare I, Halle E, Gantenberg R, Buchholz P, Jacobi HD, Presber W, Schönian G. Epidemiological study of an Acinetobacter baumannii outbreak by using polymerase chain reaction fingerprinting. J Clin Microbiol. 1993 Sep;31(9):2417–2420. [PMC free article] [PubMed]
  • Hartstein AI, Rashad AL, Liebler JM, Actis LA, Freeman J, Rourke JW, Jr, Stibolt TB, Tolmasky ME, Ellis GR, Crosa JH. Multiple intensive care unit outbreak of Acinetobacter calcoaceticus subspecies anitratus respiratory infection and colonization associated with contaminated, reusable ventilator circuits and resuscitation bags. Am J Med. 1988 Nov;85(5):624–631. [PubMed]
  • Hoffmann S, Mabeck CE, Vejlsgaard R. Bacteriuria caused by Acinetobacter calcoaceticus biovars in a normal population and in general practice. J Clin Microbiol. 1982 Sep;16(3):443–451. [PMC free article] [PubMed]
  • Horrevorts A, Bergman K, Kollée L, Breuker I, Tjernberg I, Dijkshoorn L. Clinical and epidemiological investigations of Acinetobacter genomospecies 3 in a neonatal intensive care unit. J Clin Microbiol. 1995 Jun;33(6):1567–1572. [PMC free article] [PubMed]
  • Hunter PR, Gaston MA. Numerical index of the discriminatory ability of typing systems: an application of Simpson's index of diversity. J Clin Microbiol. 1988 Nov;26(11):2465–2466. [PMC free article] [PubMed]
  • Joly-Guillou ML, Bergogne-Berezin E, Vieu JF. A study of the relationships between antibiotic resistance phenotypes, phage-typing and biotyping of 117 clinical isolates of Acinetobacter spp. J Hosp Infect. 1990 Jul;16(1):49–58. [PubMed]
  • Patterson JE, Vecchio J, Pantelick EL, Farrel P, Mazon D, Zervos MJ, Hierholzer WJ., Jr Association of contaminated gloves with transmission of Acinetobacter calcoaceticus var. anitratus in an intensive care unit. Am J Med. 1991 Nov;91(5):479–483. [PubMed]
  • Reboli AC, Houston ED, Monteforte JS, Wood CA, Hamill RJ. Discrimination of epidemic and sporadic isolates of Acinetobacter baumannii by repetitive element PCR-mediated DNA fingerprinting. J Clin Microbiol. 1994 Nov;32(11):2635–2640. [PMC free article] [PubMed]
  • Schmid J, Voss E, Soll DR. Computer-assisted methods for assessing strain relatedness in Candida albicans by fingerprinting with the moderately repetitive sequence Ca3. J Clin Microbiol. 1990 Jun;28(6):1236–1243. [PMC free article] [PubMed]
  • Seifert H, Baginski R, Schulze A, Pulverer G. Antimicrobial susceptibility of Acinetobacter species. Antimicrob Agents Chemother. 1993 Apr;37(4):750–753. [PMC free article] [PubMed]
  • Seifert H, Schulze A, Baginski R, Pulverer G. Plasmid DNA fingerprinting of Acinetobacter species other than Acinetobacter baumannii. J Clin Microbiol. 1994 Jan;32(1):82–86. [PMC free article] [PubMed]
  • Siegman-Igra Y, Bar-Yosef S, Gorea A, Avram J. Nosocomial acinetobacter meningitis secondary to invasive procedures: report of 25 cases and review. Clin Infect Dis. 1993 Nov;17(5):843–849. [PubMed]
  • Stern MJ, Ames GF, Smith NH, Robinson EC, Higgins CF. Repetitive extragenic palindromic sequences: a major component of the bacterial genome. Cell. 1984 Jul;37(3):1015–1026. [PubMed]
  • Struelens MJ, Carlier E, Maes N, Serruys E, Quint WG, van Belkum A. Nosocomial colonization and infection with multiresistant Acinetobacter baumannii: outbreak delineation using DNA macrorestriction analysis and PCR-fingerprinting. J Hosp Infect. 1993 Sep;25(1):15–32. [PubMed]
  • Tjernberg I, Ursing J. Clinical strains of Acinetobacter classified by DNA-DNA hybridization. APMIS. 1989 Jul;97(7):595–605. [PubMed]
  • Vandenbroucke-Grauls CM, Kerver AJ, Rommes JH, Jansen R, den Dekker C, Verhoef J. Endemic Acinetobacter anitratus in a surgical intensive care unit: mechanical ventilators as reservoir. Eur J Clin Microbiol Infect Dis. 1988 Aug;7(4):485–489. [PubMed]
  • Versalovic J, Koeuth T, Lupski JR. Distribution of repetitive DNA sequences in eubacteria and application to fingerprinting of bacterial genomes. Nucleic Acids Res. 1991 Dec 25;19(24):6823–6831. [PMC free article] [PubMed]
  • Woods CR, Jr, Versalovic J, Koeuth T, Lupski JR. Analysis of relationships among isolates of Citrobacter diversus by using DNA fingerprints generated by repetitive sequence-based primers in the polymerase chain reaction. J Clin Microbiol. 1992 Nov;30(11):2921–2929. [PMC free article] [PubMed]
  • Woods CR, Versalovic J, Koeuth T, Lupski JR. Whole-cell repetitive element sequence-based polymerase chain reaction allows rapid assessment of clonal relationships of bacterial isolates. J Clin Microbiol. 1993 Jul;31(7):1927–1931. [PMC free article] [PubMed]

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