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. 2001 Sep;39(9):3099-103.
doi: 10.1128/JCM.39.9.3099-3103.2001.

Identification of clinical staphylococcal isolates from humans by internal transcribed spacer PCR

I Couto  1 , S Pereira, M Miragaia, I S Sanches, H de Lencastre
Affiliations

Identification of clinical staphylococcal isolates from humans by internal transcribed spacer PCR

I Couto et al. J Clin Microbiol. 2001 Sep.

Abstract

The emergence of coagulase-negative staphylococci not only as human pathogens but also as reservoirs of antibiotic resistance determinants requires the deployment and development of methods for their rapid and reliable identification. Internal transcribed spacer-PCR (ITS-PCR) was used to identify a collection of 617 clinical staphylococcal isolates. The amplicons were resolved in high-resolution agarose gels and visually compared with the patterns obtained for the control strains of 29 staphylococcal species. Of the 617 isolates studied, 592 (95.95%) were identified by ITS-PCR and included 11 species: 302 isolates of Staphylococcus epidermidis, 157 of S. haemolyticus, 79 of S. aureus, 21 of S. hominis, 14 of S. saprophyticus, 8 of S. warneri, 6 of S. simulans, 2 of S. lugdunensis, and 1 each of S. caprae, S. carnosus, and S. cohnii. All species analyzed had unique ITS-PCR patterns, although some were very similar, namely, the group S. saprophyticus, S. cohnii, S. gallinarum, S. xylosus, S. lentus, S. equorum, and S. chromogenes, the pair S. schleiferi and S. vitulus, and the pair S. piscifermentans and S. carnosus. Four species, S. aureus, S. caprae, S. haemolyticus, and S. lugdunensis, showed polymorphisms on their ITS-PCR patterns. ITS-PCR proved to be a valuable alternative for the identification of staphylococci, offering, within the same response time and at lower cost, higher reliability than the currently available commercial systems.

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Figures

FIG. 1
FIG. 1
ITS-PCR amplification patterns for staphylococcal control strains. (A) Lanes 1 and 30, 100-bp ladder molecular size markers. Lane 2, S. auricularis; lane 3, S. vitulus; lane 4, S. schleiferi subsp. schleiferi; lane 5, S. haemolyticus; lane 6, S. capitis; lane 7, S. caprae; lane 8, S. warneri; lane 9, S. epidermidis; lane 10, S. hyicus subsp. hyicus; lane 11, S. delphini; lane 12, S. felis; lane 13, S. kloosii; lane 14, S. hominis; lane 15, S. simulans; lane 16, S. carnosus; lane 17, S. piscifermentans; lane 18, S. sciuri subsp. sciuri; lane 19, S. intermedius; lane 20, S. aureus; lane 21, S. xylosus; lane 22, S. lentus; lane 23, S. saprophyticus; lane 24, S. equorum; lane 25, S. chromogenes; lane 26, S. cohnii subsp. cohnii; lane 27, S. gallinarum; lane 28, S. arlettae; lane 29, S. lugdunensis; lane 31, S. pasteuri. (B) Lanes 1 and 9, 100-bp ladder molecular size markers. Lane 2, S. cohnii subsp. cohnii; lane 3, S. cohnii subsp. urealyticum; lane 4, S. schleiferi subsp. schleiferi; lane 5, S. schleiferi subsp. coagulans; lane 6, S. sciuri subsp. sciuri; lane 7, S. sciuri subsp. rodentium; lane 8, S. sciuri subsp. carnaticus.
FIG. 2
FIG. 2
ITS-PCR amplification patterns of clinical strains. Lanes 1 and 30, 100-bp ladder molecular size markers. Lane 2, S. epidermidis type strain; lanes 3 and 4, S. epidermidis clinical isolates; lane 5, S. haemolyticus type strain; lanes 6 and 7, S. haemolyticus clinical isolates; lane 8, S. aureus control strain; lanes 9 and 10, S. aureus clinical isolates; lane 11, S. hominis type strain; lanes 12 and 13, S. hominis clinical isolates; lane 14, S. saprophyticus type strain; lanes 15 and 16, S. saprophyticus clinical isolates; lane 17, S. warneri type strain; lanes 18 and 19, S. warneri clinical isolates; lane 20, S. simulans type strain; lane 21, S. simulans clinical isolate; lane 22, S. lugdunensis type strain; lane 23, S. lugdunensis clinical isolate; lane 24, S. caprae type strain; lane 25, S. caprae clinical isolate; lane 26, S. carnosus type strain; lane 27, S. carnosus clinical isolate; lane 28, S. cohnii subsp. cohnii type strain; lane 29, S. cohnii clinical isolate.
FIG. 3
FIG. 3
Examples of intraspecific ITS-PCR polymorphisms detected in this work. Lane 1, 100-bp ladder molecular size marker. Lanes 2 to 6, S. aureus strains; lanes 7 to 10, S. haemolyticus strains; lanes 11 and 12, S. lugdunensis strains; lanes 13 and 14, S. caprae strains. For each species, the leftmost sample is the control strain.
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