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Mechanisms of Resistance

Distribution of Insertion Sequence-Like Element IS1272 and Its Position Relative to Methicillin Resistance Genes in Clinically Important Staphylococci

Nobumichi Kobayashi, Shozo Urasawa, Nobuyuki Uehara, Naoki Watanabe
Nobumichi Kobayashi
Department of Hygieneand
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Shozo Urasawa
Department of Hygieneand
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Nobuyuki Uehara
Department of Laboratory Diagnosis,Sapporo Medical University School of Medicine, Chuo-ku, Sapporo 060-8556, Japan
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Naoki Watanabe
Department of Laboratory Diagnosis,Sapporo Medical University School of Medicine, Chuo-ku, Sapporo 060-8556, Japan
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DOI: 10.1128/AAC.43.11.2780
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ABSTRACT

The distribution of insertion sequence-like element IS1272 was analyzed for clinical isolates ofStaphylococcus aureus, Staphylococcus epidermidis, and Staphylococcus haemolyticus. In each of the staphylococcal species, IS1272 was detected in both methicillin-resistant (MR) and methicillin-susceptible strains of different genetic types. In MR isolates, IS1272 was generally located downstream of the truncated mecR1 gene (ΔmecR1), with an identical junction sequence occurring between ΔmecR1 and IS1272, although insertion of an additional gene sequence in the junction sequence was detected in one S. epidermidis isolate. These findings suggest that themec element with the rearranged form of mecR1(ΔmecR1-IS1272) has been transmitted to multiple clones of staphylococci.

Methicillin resistance of staphylococci is defined by production of penicillin-binding protein 2a (5, 15), encoded by the mecA gene, which is located on the bacterial chromosome (13). Expression ofmecA is primarily controlled by regulator elements MecR1 and MecI, encoded by mecR1 and mecI, which are located upstream of mecA (6). Although isolates of methicillin-resistant Staphylococcus aureus (MRSA) with intact mec regulatory genes show inducible resistance to beta-lactams, most recent isolates of MRSA are constitutively resistant to methicillin and have changes in the mec regulatory region (7, 8). Indeed, point mutations in the mecI ormecA operator region which are believed to have inactivated MecI were detected (7, 11, 14).

Another genomic variation, the deletion of mecI accompanied by a partial deletion of mecR1, has been reported (1, 10, 14). Archer and coworkers analyzed the MRSA strain COL, which lacks mecI and the 3′ half of mecR1, and described the presence of the insertion sequence-like element IS1272 adjacent to the incomplete mecR1 gene (1). IS1272 contains two open reading frames (ORFs) and an inverted repeat (IR) comprising 16 bases at each terminus (IRR and IRL) (2). The truncated mecR1 gene (ΔmecR1) adjacent to IS1272 has also been observed in Staphylococcus epidermidis andStaphylococcus haemolyticus; furthermore, thismecR1 deletion junction sequence was the same in S. aureus and S. epidermidis strains, as well as in anS. haemolyticus strain, Y176 (1, 2). These observations suggested that an IS1272-mediatedmecR1 deletion event may have occurred in coagulase-negative staphylococci, with subsequent horizontal transfer of the rearranged region to S. aureus.

However, it is not clear whether there are other rearranged forms of ΔmecR1 and IS1272 different from the one detected in strain COL or whether the COL-type mec DNA was transmitted from coagulase-negative staphylococci into a single clone or multiple clones of S. aureus. To investigate these points, a total of 176 clinical isolates of staphylococci from different patients admitted to the Sapporo Medical University Hospital, Sapporo, Japan, during the period from 1993 to 1998 (99 S. aureus isolates, 59 S. epidermidis isolates, and 18S. haemolyticus isolates) were analyzed in this study.S. aureus isolates of various coagulase types were selected for the present study. In addition, we analyzed three Japanese MRSA strains, MR108, MR6, and JO18, possessing truncated mecR1genes (14), which were provided by the Culture Collection Laboratory of the Institute of Medical Science at the University of Tokyo, Tokyo, Japan, and five type strains of S. haemolyticus (ATCC 15796, ATCC 29968, ATCC 29969, ATCC 29970, and ATCC 43253).

The presence of mecA, mecI, and the 5′ and 3′ halves of mecR1 was examined by PCR as described previously (9, 10). Based on the results, staphylococcal strains were classified into four groups: type 1 methicillin-resistant (MR) strains, which have both mecR1 and mecI (type 1mec region); type 2 MR strains, which lack mecIand the 3′ half of mecR1 (type 2 mec region); type 3 MR strains, lacking both mec regulator genes (type 3mec region); and methicillin-susceptible (MS) strains possessing neither the mecA nor mec regulator gene.

Detection of IS1272 was performed by PCR with two primer pairs that amplify different portions of IS1272 ORFs. IS1272 was detected in a total of 52 MR and MS isolates of the three staphylococcal species (Table1). In S. aureus, IS1272 was found in all 10 type 2 MRSA and four MSS. aureus (MSSA) isolates, while it was not detected in isolates of other mec region types. Similarly, in S. epidermidis, all of the type 2 and some of the MS isolates possessed IS1272. However, some isolates of type 1 and type 3 MR S. epidermidis were also IS1272 positive. IS1272 was detected in all of the MR and MS S. haemolyticus isolates, regardless of the mec region type, except for a type 1 isolate (SH339) and an MS strain (ATCC 29968).

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Table 1.

Detection of IS1272 in staphylococci with different mecA and mec regulator gene statuses

S. aureus isolates were genetically classified by protein A typing, which measures the number of 24-bp repeating units in theXr region of the protein A gene, as described previously (4, 12). As shown in Table 2, IS1272 was found in coagulase type III, IV, and VII MRSA with different protein A types (repeat no. 5, 8, 9, 10, 11, and 12). The three MRSA strains not from the Sapporo Medical University Hospital—MR108, MR6, and JO18—possessed IS1272 and showed identical coagulase (IV) and protein A (repeat no. 9) types. IS1272 was detected in coagulase type IV, V, and VII MSSA isolates with different protein A types. In contrast, no coagulase type II MRSA and MSSA possessed this genetic element.

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Table 2.

Distribution of IS1272 in S. aureusstrains of various coagulase and protein A types

S. epidermidis and S. haemolyticus were typed by arbitrarily primed PCR fingerprinting with ERIC2 primer (16) and M13 reverse primer (3) as described previously (16). As a result, S. epidermidis andS. haemolyticus carrying IS1272 were differentiated into seven and two genetic types, respectively (data not shown).

The presence of the junction sequence between ΔmecR1 and IS1272 was examined for all of the type 2 isolates by PCR with primer pair mecRA1 (10) and mDA2 (5′-GATGTCTGTCGAGGACTC-3′), which are complementary to the sequences in mecR1 and IS1272, respectively. A PCR product of 1,287 bp, which was expected from the sequence ofS. aureus COL (GenBank accession no. L14017), was obtained for 22 isolates, although no PCR products were generated for the two MR S. haemolyticus isolates with a type 2mec region. However, a PCR product which was approximately 1,400 bp longer than those from other isolates was generated for oneS. epidermidis strain (SH194).

The nucleotide sequence between the 3′ end of ΔmecR1 and the IRL of IS1272 was determined directly from these PCR products by the dideoxynucleotide chain termination method (12). The 3′ ends of the mecR1 genes of the 23 type 2 isolates were identical to those of S. aureus COL andS. haemolyticus Y176 (1, 2) (GenBank accession no. L14017 and U35635, respectively), containing a 9-base divergent sequence not found in mecR1 (Fig.1a). The truncated mecR1 had an ORF of 984 bp and was suggested to encode an incomplete MecR1 product comprising 328 amino acids. The ΔmecR1-IS1272 junction sequences of all isolates except for SH194 consisted of 248 bases, a length almost identical to those of strains COL and Y176. However, compared with published sequences of these strains, two additional nucleotides, T and A, were identified in all staphylococcal genomes examined (Fig. 1a). Furthermore, a single nucleotide was substituted in the junction sequence of S. aureus SH475 (Fig. 1a).

Fig. 1.
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Fig. 1.

(a) Nucleotide sequences of the 3′-end portion of ΔmecR1, the ΔmecR1-IS1272 junction sequence (numbered on the sequence from 1 to 248), and the IRL side of IS1272 determined for staphylococci in the present study. Termination codons for ΔmecR1 and ORF2 of IS1272 are shown in boldface type and are followed by vertical lines. Dotted underlining indicates divergent terminal sequence of ΔmecR1, while solid underlining shows the IRLs. Lines above the nucleotide sequences TT and AAAAAA, under solid inverted triangles, denote the regions where an additional T and A were identified, respectively, compared with those of strains COL and Y176 (1, 2). A single nucleotide in the junction sequence which was substituted in S. aureusSH475 is boxed. (b) In S. epidermidis SH194, an additional gene sequence (approximately 1,400 bp) was found in the IRL-side terminal portion of the junction sequence. Lowercase letters indicate partial nucleotide sequences at both ends of the inserted sequence, which initiates with adenine (*1) after the 247th nucleotide of the junction sequence (a) and terminates at the guanine (*2) close to the IRL of IS1272 (b).

In S. epidermidis SH194, the truncated portion ofmecR1, the ΔmecR1-side 247-base junction sequence (including two additional bases), and the IRL of IS1272 were identical to those of most other isolates. However, an insertion of additional DNA was found in the junction sequence at the site close to the IRL. Partial sequences of both sides of the additional DNA are shown in Fig. 1.

In the present study, IS1272 was found in S. aureus, S. epidermidis, and S. haemolyticus strains with the type 2 mec region, together with almost identical ΔmecR1-IS1272junction sequences. This finding supports the hypothesis that amec element with ΔmecR1 and IS1272has been transmitted horizontally among staphylococci. However, IS1272 was also detected in type 1 and type 3 staphylococci and in MSSA in the present study. Hence, IS1272 may have been disseminated among staphylococci irrespective of the presence ofmec DNA, although this genetic element appears to have been originally resident in S. haemolyticus (2).

Molecular epidemiologic typing of S. aureus indicated that truncated mecR1 and IS1272 are distributed inS. aureus strains of different coagulase and protein A types and also in S. epidermidis and S. haemolyticusstrains with some distinct genetic types. These findings suggested that the mec element containing ΔmecR1 and IS1272 might have been transmitted to multiple clones of staphylococci.

It is notable that an additional large DNA sequence is inserted in the IRL-side terminal portion of the ΔmecR1-IS1272junction sequence in S. epidermidis SH194, which exhibited a novel rearrangement of ΔmecR1 with IS1272. Furthermore, the ΔmecR1-IS1272 junction sequence was not detected in two isolates of type 2 MR S. haemolyticus, suggesting the presence of an unknown form of deletion junction sequence in mec regulator genes. Thus,mec regulator genes may contain various genomic changes, which seem to have occurred multiple times in coagulase-negative staphylococci.

FOOTNOTES

    • Received 14 May 1999.
    • Returned for modification 29 June 1999.
    • Accepted 18 August 1999.
  • Copyright © 1999 American Society for Microbiology

REFERENCES

  1. 1.↵
    1. Archer G. L.,
    2. Niemeyer D. M.,
    3. Thanassi J. A.,
    4. Pucci M. J.
    Dissemination among staphylococci of DNA sequences associated with methicillin resistance. Antimicrob. Agents Chemother. 38 1994 447 454
    OpenUrlAbstract/FREE Full Text
  2. 2.↵
    1. Archer G. L.,
    2. Thanassi J. A.,
    3. Niemeyer D. M.,
    4. Pucci M. J.
    Characterization of IS1272, an insertion sequence-like element from Staphylococcus haemolyticus. Antimicrob. Agents Chemother. 40 1996 924 929
    OpenUrlAbstract/FREE Full Text
  3. 3.↵
    1. Fang F. C.,
    2. McClelland M.,
    3. Guiney D. G.,
    4. Jackson M. M.,
    5. Hartstein A. I.,
    6. Morthland V. H.,
    7. Davis C. E.,
    8. McPherson D. C.,
    9. Welsh J.
    Value of molecular epidemiologic analysis in a nosocomial methicillin-resistant Staphylococcus aureus outbreak. JAMA 270 1993 1323 1328
    OpenUrlCrossRefPubMed
  4. 4.↵
    1. Frénay H. M. E.,
    2. Theelen J. P. G.,
    3. Schouls L. M.,
    4. Vandenbroucke-Grauls C. M. J. E.,
    5. Verhoef J.,
    6. van Leeuwen W. J.,
    7. Mooi F. R.
    Discrimination of epidemic and nonepidemic methicillin-resistant Staphylococcus aureus strains on the basis of protein A gene polymorphism. J. Clin. Microbiol. 32 1994 846 847
    OpenUrlAbstract/FREE Full Text
  5. 5.↵
    1. Hartman B. J.,
    2. Tomasz A.
    Low-affinity penicillin-binding protein associated with β-lactam resistance in Staphylococcus aureus. J. Bacteriol. 158 1984 513 516
    OpenUrlAbstract/FREE Full Text
  6. 6.↵
    1. Hiramatsu K.,
    2. Asada K.,
    3. Suzuki E.,
    4. Okonogi K.,
    5. Yokota T.
    Molecular cloning and nucleotide sequence determination of the regulator region of mecA gene in methicillin-resistant Staphylococcus aureus (MRSA). FEBS Lett. 298 1992 133 136
    OpenUrlCrossRefPubMedWeb of Science
  7. 7.↵
    1. Hiramatsu K.
    Molecular evolution of MRSA. Microbiol. Immunol. 39 1995 531 543
    OpenUrlCrossRefPubMedWeb of Science
  8. 8.↵
    1. Hiramatsu K.,
    2. Kondo N.,
    3. Ito T.
    Genetic basis for molecular epidemiology of MRSA. J. Infect. Chemother. 2 1996 117 129
    OpenUrlCrossRef
  9. 9.↵
    1. Kobayashi N.,
    2. Wu H.,
    3. Kojima K.,
    4. Taniguchi K.,
    5. Urasawa S.,
    6. Uehara N.,
    7. Omizu Y.,
    8. Kishi Y.,
    9. Yagihashi A.,
    10. Kurokawa I.
    Detection of mecA,femA, and femB genes in clinical strains of staphylococci using polymerase chain reaction. Epidemiol. Infect. 113 1994 259 266
    OpenUrlCrossRefPubMedWeb of Science
  10. 10.↵
    1. Kobayashi N.,
    2. Taniguchi K.,
    3. Kojima K.,
    4. Urasawa S.,
    5. Uehara N.,
    6. Omizu Y.,
    7. Kishi Y.,
    8. Yagihashi A.,
    9. Kurokawa I.,
    10. Watanabe N.
    Genomic diversity of mec regulator genes in methicillin-resistant Staphylococcus aureus and Staphylococcus epidermidis. Epidemiol. Infect. 117 1996 289 295
    OpenUrlCrossRefPubMed
  11. 11.↵
    1. Kobayashi N.,
    2. Taniguchi K.,
    3. Urasawa S.
    Analysis of diversity of mutations in the mecI gene and mecA promoter/operator region of methicillin-resistant Staphylococcus aureus and Staphylococcus epidermidis. Antimicrob. Agents Chemother. 42 1998 717 720
    OpenUrlAbstract/FREE Full Text
  12. 12.↵
    1. Kobayashi N.,
    2. Urasawa S.,
    3. Uehara N.,
    4. Watanabe N.
    Analysis of genomic diversity within the Xr-region of the protein A gene in clinical isolates of Staphylococcus aureus. Epidemiol. Infect. 122 1999 241 249
    OpenUrlCrossRefPubMed
  13. 13.↵
    1. Song M. D.,
    2. Wachi M.,
    3. Doi M.,
    4. Ishino F.,
    5. Matsuhashi M.
    Evolution of an inducible penicillin-target protein in methicillin-resistant Staphylococcus aureus by gene fusion. FEBS Lett. 221 1987 167 171
    OpenUrlCrossRefPubMedWeb of Science
  14. 14.↵
    1. Suzuki E.,
    2. Kuwahara-Arai K.,
    3. Richardson J. F.,
    4. Hiramatsu K.
    Distribution of mec regulator genes in methicillin-resistant Staphylococcus clinical strains. Antimicrob. Agents Chemother. 37 1993 1219 1226
    OpenUrlAbstract/FREE Full Text
  15. 15.↵
    1. Utsui Y.,
    2. Yokota T.
    Role of an altered penicillin-binding protein in methicillin- and cephem-resistant Staphylococcus aureus. Antimicrob. Agents Chemother. 28 1985 397 403
    OpenUrlAbstract/FREE Full Text
  16. 16.↵
    1. van Belkum A.,
    2. Bax R.,
    3. Peerbooms P.,
    4. Goessens W. H. F.,
    5. Van Leeuwen N.,
    6. Quint W. G. V.
    Comparison of phage typing and DNA fingerprinting by polymerase chain reaction for discrimination of methicillin-resistant Staphylococcus aureus strains. J. Clin. Microbiol. 31 1993 798 803
    OpenUrlAbstract/FREE Full Text
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Distribution of Insertion Sequence-Like Element IS1272 and Its Position Relative to Methicillin Resistance Genes in Clinically Important Staphylococci
Nobumichi Kobayashi, Shozo Urasawa, Nobuyuki Uehara, Naoki Watanabe
Antimicrobial Agents and Chemotherapy Nov 1999, 43 (11) 2780-2782; DOI: 10.1128/AAC.43.11.2780

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Distribution of Insertion Sequence-Like Element IS1272 and Its Position Relative to Methicillin Resistance Genes in Clinically Important Staphylococci
Nobumichi Kobayashi, Shozo Urasawa, Nobuyuki Uehara, Naoki Watanabe
Antimicrobial Agents and Chemotherapy Nov 1999, 43 (11) 2780-2782; DOI: 10.1128/AAC.43.11.2780
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