Genetic organization of the mecA region in methicillin-susceptible and methicillin-resistant strains of Staphylococcus sciuri

Abstract

A homolog of the Staphylococcus aureus methicillin resistance gene mecA was recently shown to be ubiquitous in independent isolates of the animal species Staphylococcus sciuri. The mecA gene homolog and regions flanking it were cloned and sequenced from four strains of S. sciuri: strain K1 (ATCC 29062), a representative of S. sciuri subsp. sciuri; two strains (K3 and K8) representing S. sciuri subsp. rodentius; and strain K11, a representative of S. sciuri subsp. carnaticum. Strains K1 and K11 were susceptible to methicillin, while strains K3 and K8 showed heterogeneous resistance. The mecA genes of strains K1 and K11 and one of the two copies of mecA (mecA1) present in strain K3 had virtually identical DNA sequences in the mecA gene and were similar in genetic organization in the flanking regions. In contrast, the single copy of mecA in strain K8 and the second copy of mecA (mecA2) in strain K3 had mecA DNA sequences identical to that of S. aureus mecA, and the mecA region in these two strains was also similar to that of the same region in the S. aureus strain used for comparison. Interestingly, an open reading frame defining an N-terminal truncated polypeptide, NTORF101, with a high degree of homology to a DNA segment in the hypervariable region of methicillin-resistant S. aureus (and also similar to the Escherichia coli gene ugpQ) was also identified downstream of the mecA homolog of strain K11, representing S. sciuri subsp. carnaticum. The ugpQ-like gene is not present in methicillin-susceptible strains of S. aureus. The presence of such a ugpQ-like gene together with the homolog of mecA in strain K11 supports the speculation that these genetic elements may be evolutionary relatives and/or precursors of the genetic determinant of methicillin resistance in S. aureus.

FIG. 1

MseI fingerprints of the mecA genes from a selected group of S. sciuri strains. Lanes 1 and 15, pBR322 digested with HpaII size markers; lanes 2 and 3, fingerprints of S. sciuri K1 and S. sciuri subsp. carnaticum K11 (pattern I); lanes 4 and 5, fingerprints of S. sciuri K47 and K105 (pattern II); lanes 6 and 7, fingerprints of S. sciuri K2 and S. sciuri subsp. carnaticum K61 (pattern III); lanes 8 and 9, fingerprints of S. sciuri K165 and KL064 (pattern IV); lanes 10 and 11, fingerprints of S. sciuri subsp. rodentius K3 (mecA1) and S. sciuri subsp. carnaticum K30 (pattern V); lanes 12 to 14, fingerprints of S. sciuri subsp. rodentius K3 (mecA2) and K8 and S. aureus COL (pattern VI).

FIG. 2

Genetic organization of the mecA regions of S. sciuri (K1), S. sciuri subsp. rodentius (K8 and K3), S. sciuri subsp. carnaticum (K11), and S. aureus. The S. aureus mecA region (c) was assembled by using DNA sequences from the database. The direction of gene transcription is shown by the arrows. Identical symbols are used to indicate sequences of significant similarity. Dashed symbols indicate regions determined by PCR analysis. Homologies between short DNA segments are indicated by • (near ORF454 of K3 and near ORF142 of S. aureus) or by •• (near CTORF225 of K3 and near CTORF168 of S. aureus). Accession numbers for these sequences are as follows: K8 (5,596 bp), Y13096; K3 mecA2 (6,368 bp), Y13095; K11 (6,684 bp), Y13094; K3 mecA1 (5,806 bp), Y13052; and K1 (5,068 bp), Y09223. To facilitate comparison, relevant DNA sequences of the mecA region of S. aureus are also provided: mecRI (1) (accession no. L14020), mecA (23) (accession no. X52593), hypervariable region (22) (accession no. X52594), and IS431 (4, 5) (accession no. X53818). These sequences were assembled to provide a map for the entire S. aureus mecA and flanking region (c [9,047 bp; accession no. Y14051]). Overlaps or gaps at junctions in this map were determined by sequencing the appropriate PCR products of S. aureus BB589 (7).

FIG. 3

DNA sequence alignment of short segments in the area outside the mecA coding region. (a) DNA sequence at the N terminus of CTORF261 in K3 mecA2 compared with the corresponding region in S. aureus. Bold letters represent the direct repeat which is the likely position for IS insertion. (b) DNA sequence of NTORF101 in K11 compared with the intervening area between ORF142 and ORF145 in S. aureus. The nucleotide at position 6239 (C) in the S. aureus sequence was replaced with an A residue, creating an extra stop codon and causing interruption of the UgpQ-like polypeptide.

FIG. 4

Relationship of the E. coli UgpQ (ECUGPQ) and S. aureus hypervariable regions in ORF145 (SAORF145) and ORF44 (SAORF44) and NTORF101 of strain K11 (K11NTORF101). (a) ORF145 was 57% homologous to the N-terminal part of E. coli UgpQ. (b) ORF44 was 65% similar to the C-terminal part of UgpQ. (c) NTORF101 of strain K11 showed 58% similarity to the C terminus of E. coli UgpQ. (d) Multisequence alignment of S. aureus ORF44, K11 NTORF101, and K8 NTORF101, documenting a high degree of similarity.