Antimicrobial resistance gene shuffling and a three-element mobilisation system in the monophasic Salmonella typhimurium strain ST1030.

In this study we describe the genetic elements and the antimicrobial resistance units (RUs) harboured by the Salmonella Typhimurium monophasic variant 1,4,[5],12:i:- strain ST1030. Of the three identified RUs two were chromosomal, RU1 (IS26-blaTEM-1-IS26-strAB-sul2- IS26) and RU2 (IS26-tetR(B)-tetA(B)-ΔIS26), and one, RU3 (a sul3-associated class 1 integron with cassette array dfrA12-orfF-aadA2-cmlA1-aadA1), was embedded in a Tn21-derived element harboured by the conjugative I1 plasmid pST1030-1A. IS26 elements mediated the antimicrobial resistance gene (ARG) shuffling and this gave rise to pST1030-1A derivatives with different sets of ARGs. ST1030 also harboured two ColE1-like plasmids of which one, pST1030-2A, was mobilisable and the target of an intracellular translocation of the Tn21-derived element; the second (pST1030-3) was an orphan mob-associated oriT plasmid co-transferred with pST1030-1A and pST1030-2A. pST1030-2A and pST1030-3 also carried a parA gene and a type III restriction modification system, respectively. Overall analysis of our data reinforces the role played by IS26, Tn21-derived elements and non-conjugative plasmids in the spread of ARGs and supplies the first evidence, at least in Salmonella, for the identification of a natural isolate harbouring a three-element mobilisation system in the same cell.

IS26 mediated antimicrobial resistance gene shuffling from the chromosome to a mosaic conjugative FII plasmid.

In the present study we report the identification of a sul3-associated class 1 integron containing the dfrA12-orfF-aadA2-cmlA1-aadA1-qacH array embedded in a Tn21-derived element that is part of a conjugative FII plasmid named pST1007-1A. The plasmid was identified in the Salmonella Typhimurium strain ST1007, a member of a clinically relevant clonal MDR lineage diffuse in Italy. ST1007 exhibited resistance to ampicillin, chloramphenicol, streptomycin, sulphamethoxazole, tetracycline and trimethoprim encoded by blaTEM-1, cmlA1, (aadA1, aadA2, strAB), (sul2, sul3), tet(B) and dfrA12 genes, respectively. Apart from pST1007-1A, ST1007 also harbours two chromosome-integrated resistance units RU1 (blaTEM-1-sul2-strAB) and RU2 (tet(B)), flanked by IS26 elements. RU1 and RU2 were able to move as translocatable units, respectively TU1 and TU2, and integrate via IS26 mediated recombination into pST1007-1A. A family of conjugative plasmids, harbouring different sets of antimicrobial resistance genes (ARG) was then generated: pST1007-1B (dfrA12-aadA2-cmlA1-aadA1-sul3- tet(B)), pST1007-1C (dfrA12-aadA2-cmlA1-aadA1-sul3-blaTEM-1-sul2-strAB), pST1007-1D (blaTEM-1-sul2-strAB), pST1007-1E (tet(B)) and pST1007-1F (dfrA12-aadA2-cmlA1-aadA1-sul3- tet(B) -blaTEM-1-sul2-strAB). pST1007-1A is also a mosaic plasmid containing two distinct DNA fragments acquired from I1 plasmids through recombination within the repA4, rfsF and repeat-3 sites. This study further highlights the role played by IS26 in intracellular ARGs shuffling. Moreover, attention has been focused on recombination hot spots that might play a key role in generating mosaic plasmids.

A novel group of IncQ1 plasmids conferring multidrug resistance.

The IncQ is a group of non-conjugative but mobilisable plasmids that are found and stably maintained in a wide range of bacteria contributing to the spread of antimicrobial resistance genes and to the insurgence of multidrug resistant bacteria. Here we report the identification, in clinical Salmonella Typhimurium strains, of an IncQ1 plasmid (pNUC) which confers resistance to sulfamethoxazole, streptomycin and tetracycline through the presence of sul2, strAB and tetA genes, respectively. pNUC was detected in five multidrug resistant S. Typhimurium strains collected in Southern Italy from various hospitals and years of isolation. Bioinformatics analyses highlighted the presence of pNUC-like plasmids in pathogenic bacteria of various Enterobacteriaceae genera or species. Taken as a whole, these plasmids constitute a novel group of IncQ1 plasmids that might have originated through recombination events between a tetR-tetA gene cluster (possibly derived from a Tn1721) and a recipient IncQ1 plasmid related to RSF1010. Our findings raise concerns regarding the possible contribution of the newly identified group of IncQ1 plasmids to the spread of tetracycline resistance.

SXT-related integrating conjugative element and IncC plasmids in Vibrio cholerae O1 strains in Eastern Africa.

OBJECTIVES: The objective of this study was to investigate the extent of resistance patterns and associated mobile genetic elements in epidemic V. cholerae O1 El Tor strains isolated from Eastern Africa in the late 1990s. METHODS: Self-transmissible genetic elements and associated clusters of genes encoding resistance were detected by conjugation experiments. Detection of SXT-related integrating conjugative elements (ICEs) and associated antibiotic resistance genes was performed by PCR to amplify the SXT element-integrase gene (int), right SXT element-chromosome junction (attP-prfC) and genes conferring resistance to chloramphenicol (floR), sulfamethoxazole (sulII), streptomycin (strA) and trimethoprim (dfrA1). Genomic relatedness was established by random amplified polymorphic DNA patterns. RESULTS: Of 224 strains analysed, 200 isolates exhibited resistance to four or more antimicrobials. An IncC plasmid, encoding resistance to ampicillin, chloramphenicol, streptomycin, sulfamethoxazole and trimethoprim, conferred multidrug resistance to 113 strains isolated from Somalia and Ethiopia, whereas an SXT-related ICE, encoding resistance to chloramphenicol, streptomycin, sulfamethoxazole and trimethoprim, conferred multidrug resistance to 74 strains isolated from Sudan, Kenya and Tanzania. CONCLUSIONS: This study has shown the spread of SXT-related ICEs among V. cholerae O1 African isolates. It has also highlighted the role of two distinct genetic elements in conferring multiple resistance to the two distinct groups of V. cholerae O1 strains that, in the late 1990s, spread through Eastern Africa, a critical geographic region for the persistence and transmission of cholera to the entire continent.

Clonal relationship among Vibrio cholerae O1 El Tor strains causing the largest cholera epidemic in Kenya in the late 1990s.

Eighty Vibrio cholerae O1 strains selected to represent the 1998-to-1999 history of the largest cholera epidemic in Kenya were characterized by ribotyping, antimicrobial susceptibility, and random amplified polymorphic DNA patterns. Except for 19 strains from 4 local outbreaks in North Eastern Province along the Somalia border, the other 61 strains from 25 outbreaks occurring in districts scattered around the country were all ribotype B27 and resistant to chloramphenicol, spectinomycin, streptomycin, sulfamethoxazole, and trimethoprim. The 61 strains showed similar and specific amplified DNA patterns. These findings indicate that the predominant strains that caused the Kenyan epidemic had a clonal origin and suggest that ribotype B27 strains, which first appeared in West Africa in 1994, have had a rapid spread to eastern Africa.