Possible role of insertion sequence IS257 in dissemination and expression of high- and low-level trimethoprim resistance in staphylococci

Leelaporn, Amornrut; Firth, Neville; Byrne, Mary E.; Roper, E; Skurray, Ronald A.

doi:10.1128/aac.38.10.2238

Cited by 40 publications

(35 citation statements)

References 42 publications

(42 reference statements)

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“…Moreover, high-and low-level trimethoprim resistance in Staphylococcus aureus is mediated by genomic deletions adjacent to a copy of IS257 inserted upstream from the dihydrofolate reductase gene, dfrA. In this case, the insertion sequence encodes a Ϫ35 sequence that is necessary for full promoter activity (18). IS6110 can promote gene expression in M. tuberculosis 210 (3).…”

Section: Methodsmentioning

confidence: 99%

IS 6110 Mediates Increased Transcription of the phoP Virulence Gene in a Multidrug-Resistant Clinical Isolate Responsible for Tuberculosis Outbreaks

et al. 2004

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Drug resistance in Mycobacterium tuberculosis complex strains is solely due to chromosomal mutations that could affect bacterial virulence. Molecular epidemiology studies have shown that resistant strains are less likely to be clustered than susceptible strains. However, a few multidrug-resistant (MDR) M. tuberculosis complex strains have been described as causing outbreaks, suggesting that they have restored virulence or increased transmission. One of the biggest MDR tuberculosis outbreaks documented to date was caused by the B strain of M. bovis. Restriction fragment length polymorphism fingerprinting revealed that the B strain contains two copies of IS6110. Here, we mapped and sequenced the regions flanking the two copies of IS6110 in the B strain. Ligation-mediated PCR showed that one of these IS6110 copies is located within the promoter region of phoP, a transcriptional regulator that is essential for M. tuberculosis virulence. We used PCR to screen 219 MDR M. tuberculosis complex strains (90.4% of all MDR isolates) isolated in Spain between 1998 and 2002 and found that the B strain was the only strain that contained a copy of IS6110 in the phoP promoter. To determine whether IS6110 affects phoP promoter activity in the B strain, we individually cloned the phoP gene and its promoter region (including IS6110 from the B strain and the equivalent region from M. tuberculosis without IS6110 as a control) into a mycobacterial replicative plasmid and transformed M. smegmatis with the resulting plasmid. Primer extension analysis showed that phoP transcription was strongly upregulated when the promoter region contained IS6110, as in the case of the B strain.Tuberculosis (TB) is currently one of the leading causes of mortality throughout the world (8,25,27). The human immunodeficiency virus-AIDS pandemic, the deterioration of public health systems in developing countries, and the emergence of multidrug-resistant (MDR) Mycobacterium tuberculosis complex strains have further contributed to the spread of TB. Knowledge of the molecular mechanisms involved in the bacillus-host cell interaction is essential for developing adequate strategies for TB control. Recent advances in the genetic manipulation of mycobacteria (2, 29) combined with the publication of the complete M. tuberculosis genome sequence (6) have made it possible to study the contribution of individual genes to M. tuberculosis virulence (5, 7). However, little is known about the regulatory and expression mechanisms that determine the virulence of clinical isolates of M. tuberculosis.Insertion sequence (IS) 6110 has been extensively used for molecular typing of M. tuberculosis strains. Restriction fragment length polymorphism (RFLP) analysis using IS6110 as a probe is currently the most common molecular method used to type M. tuberculosis complex strains. However, the physiological role and impact of specific IS6110 insertions on the biology of bacilli are not well known. IS6110 fingerprinting studies have demonstrated heterogeneity between virulence and transm...

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Section: Methodsmentioning

confidence: 99%

IS 6110 Mediates Increased Transcription of the phoP Virulence Gene in a Multidrug-Resistant Clinical Isolate Responsible for Tuberculosis Outbreaks

et al. 2004

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“…7) (15). IS257 has been found to insert into several locations in pT181, with no apparent insertion site specificity (16,25,35).…”

Section: Discussionmentioning

confidence: 99%

“…The canonical promoter consensus is shown at the top. The TSPs mapped for dfrA on pSK1 (15) and tetA(K) on SK1660 (this study) are indicated. Eight-base pair target duplication sequences are denoted by dashed arrows.…”

Section: Discussionmentioning

confidence: 99%

“…Total cellular RNA was isolated from S. aureus strains as previously described (15). Transcript mapping was performed essentially as described by Ausubel et al (2).…”

Section: Methodsmentioning

confidence: 99%

“…In strains exhibiting highlevel trimethoprim resistance, transcription of dfrA is directed by a hybrid promoter consisting of a Ϫ35 sequence encoded within the end of IS257 and a Ϫ10 sequence located in the adjacent sequence (15). Sequence analysis has suggested that an analogous IS257-derived hybrid promoter might also be responsible for transcription of tetA(K) in the chromosomal pT181-like plasmid cointegrate of strains such as SK1660 (33).…”

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An IS 257 -Derived Hybrid Promoter Directs Transcription of a tetA (K) Tetracycline Resistance Gene in the Staphylococcus aureus Chromosomal mec Region

2000

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Transcription of the tetA(K) tetracycline resistance determinant encoded by an IS257-flanked cointegrated copy of a pT181-like plasmid, located within the chromosomal mec region of a methicillin-resistant Staphylococcus aureus isolate, has been investigated. The results demonstrated that transcription of tetA(K) in this strain is directed by both an IS257-derived hybrid promoter, which is stronger than the native tetA(K) promoter in the autonomous form of pT181, and a complete outwardly directed promoter identified within one end of IS257. Despite lower gene dosage, the chromosomal configuration was shown to afford a higher level of resistance than that mediated by pT181 in an autonomous multicopy state. Furthermore, competition studies revealed that a strain carrying the chromosomal tetA(K) determinant exhibited a higher level of fitness in the presence of tetracycline but not in its absence. This finding suggests that tetracycline has been a selective factor in the emergence of strains carrying a cointegrated pT181-like plasmid in their chromosomes. The results highlight the potential of IS257 to influence the expression of neighboring genes, a property likely to enhance its capacity to mediate advantageous genetic rearrangements.

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Bacterial Transposons—An Increasingly Diverse Group of Elements

Whittle

Salyers

2002

Modern Microbial Genetics

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Possible role of insertion sequence IS257 in dissemination and expression of high- and low-level trimethoprim resistance in staphylococci

Cited by 40 publications

References 42 publications

IS 6110 Mediates Increased Transcription of the phoP Virulence Gene in a Multidrug-Resistant Clinical Isolate Responsible for Tuberculosis Outbreaks

IS 6110 Mediates Increased Transcription of the phoP Virulence Gene in a Multidrug-Resistant Clinical Isolate Responsible for Tuberculosis Outbreaks

An IS 257 -Derived Hybrid Promoter Directs Transcription of a tetA (K) Tetracycline Resistance Gene in the Staphylococcus aureus Chromosomal mec Region

Bacterial Transposons—An Increasingly Diverse Group of Elements

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