New variants of the tet(M) gene in Clostridium difficile clinical isolates harbouring Tn916-like elements.
OBJECTIVE
To detect Tn916-like elements in Clostridium difficile clinical isolates from different time periods and to analyse the genetic structure of these elements, in particular the tet(M) region.


METHODS
Ninety C. difficile clinical isolates were examined by PCR assays for tet(M) and int, which are markers for the Tn916 family of elements. Positive isolates were typed by PCR-ribotyping, and tetracycline MIC values were evaluated by Etest. The genetic organization of the Tn916 elements was investigated by PCR mapping and hybridization assays. The tet(M) region of eight selected C. difficile isolates was sequenced.


RESULTS
Nineteen isolates were tet(M)/int positive and the majority (12/19) belonged to PCR-ribotype R, previously found to be predominant in clinical strains of more recent isolation. Eleven isolates were tetracycline resistant, three inducibly resistant and five susceptible. Fifty-eight per cent of the C. difficile isolates harboured one Tn916 element and 42% harboured two. Most isolates showed elements with a genetic organization very similar to that of Enterococcus faecalis DS16 Tn916. Sequence analysis highlighted variations in the leader peptide and six tet(M) variants were identified, five of which have never been described before.


CONCLUSIONS
C. difficile isolates harbouring Tn916-like elements have mainly been isolated since 1997, suggesting a recent circulation of these elements among C. difficile strains in Italian hospitals. Molecular analysis of these Tn916-like elements showed that they may have different genetic structures and carry new tet(M) alleles.
Department of Infectious, Parasitic and Immune-mediated Diseases, Istituto Superiore di Sanità Viale, Regina Elena 299, 00161 Rome, Italy.
Patrizia
P
Spigaglia
Fabrizio
F
Barbanti
Paola
P
Mastrantonio
D001426:Bacterial Proteins / Q000235:genetics*; D016360:Clostridioides difficile / Q000187:drug effects / Q000235:genetics* / Q000302:isolation & purification; D003428:Cross Infection / Q000382:microbiology; D004251:DNA Transposable Elements / Q000235:genetics*; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics; D004761:Enterocolitis, Pseudomembranous / Q000382:microbiology; D006801:Humans; D007558:Italy; D008826:Microbial Sensitivity Tests; D008969:Molecular Sequence Data; D009693:Nucleic Acid Hybridization; D010802:Phylogeny; D016133:Polymerase Chain Reaction; D011110:Polymorphism, Genetic; D021521:Ribotyping; D017422:Sequence Analysis, DNA; D017385:Sequence Homology; D013753:Tetracycline Resistance / Q000235:genetics*
D001426:Bacterial Proteins; D004251:DNA Transposable Elements; D004269:DNA, Bacterial; C083138:Tet M resistance protein, Bacteria