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Diagram of 14 distinct upstream flanking sequences and 10 distinct downstream flanking sequences of IS1237 in the L. xyli subsp. cynodontis strain #3 genome. The recognition sequences TAA and TTA are indicated by bold type. Translational start codons are underlined. The shaded arrows with GenBank accession numbers represent contiguous flanking ORFs. The shaded boxes indicate the truncated genes which were interrupted by IS1237. Brief descriptions of flanking ORFs, where GenBank numbers are indicated, are given to the left or right of the flanking ORFs. The spacers between IS1237 elements and start or stop codons of the flanking genes are represented by bidirectional arrows, and their lengths are indicated. ID, identity; aa, amino acids.
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We performed a comprehensive study of the distribution and function of an insertion sequence (IS) element, IS1237, in the genome of Leifsonia xyli subsp. cynodontis, a useful genetic carrier for expressing beneficial foreign genes in plants. Two shorter IS1237 isoforms, IS1237d1 and IS1237d2 resulting from precise deletion between two nonperfect re...
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... A similar situation occurs in the case of the mosaic TE generated by ISAba1 and ISAba9, described above (Figure 2f). IS1237 (IS5 family) is unusual because it contains two active outward promoters at both termini [109] (Figure 2g). These promoters differ in their activity (that at the 3 end is stronger), but both were shown to drive the transcription of adjacent genes in different Gram-positive bacteria. ...
The rapidly emerging phenomenon of antibiotic resistance threatens to substantially reduce the efficacy of available antibacterial therapies. Dissemination of resistance, even between phylogenetically distant bacterial species, is mediated mainly by mobile genetic elements, considered to be natural vectors of horizontal gene transfer. Transposable elements (TEs) play a major role in this process—due to their highly recombinogenic nature they can mobilize adjacent genes and can introduce them into the pool of mobile DNA. Studies investigating this phenomenon usually focus on the genetic load of transposons and the molecular basis of their mobility. However, genes introduced into evolutionarily distant hosts are not necessarily expressed. As a result, bacterial genomes contain a reservoir of transcriptionally silent genetic information that can be activated by various transposon-related recombination events. The TEs themselves along with processes associated with their transposition can introduce promoters into random genomic locations. Thus, similarly to integrons, they have the potential to convert dormant genes into fully functional antibiotic resistance determinants. In this review, we describe the genetic basis of such events and by extension the mechanisms promoting the emergence of new drug-resistant bacterial strains.
... Several studies have investigated the dissemination of carbapenemases such as bla NDM , bla OXA-51-like , and bla OXA-23-like in A. baumannii and have reported a prevalence of more than 70% among isolates in Egypt (12,13). The propagation of carbapenemase-encoding genes is in large part due to their association with integrons and plasmids (14,15), allowing their rapid transmission and accumulation. Insertion sequences (ISs) in the region surrounding these genes cause their overexpression by providing promoter sequences, resulting in higher MICs for carbapenems (16)(17)(18)(19)(20). ...
Acinetobacter baumannii represents a major health threat, in particular among immunocompromised cancer patients. The rise in carbapenem-resistant A. baumannii , and the development of resistance to the last-resort antimicrobial agent colistin, complicates the management of A. baumannii outbreaks and increases mortality rates.
... IS elements and their transposition have been implicated in the evolution of their hosts as they contribute to a variety of genomic rearrangements (deletions, inversions and duplications) that can result in gene inactivation and/or affect the expression of ISRme20 IS21 1977 1 ISRme4 IS21 2469 2 ISRme9 IS21 2688 1 IS1090 IS256 1343 4 ISRme11 IS3 (IS150) 1231 2 ISRme12 IS3 (IS150) 1454 1 ISRme17 IS3 (IS150) 1678 1 IS1087B IS3 (IS2) 1330 2 ISRme3 IS3 (IS3) 1288 3 5 2 ISRme15 IS3 (IS51) 1325 1 1 IS1086 IS30 1106 1 1 1 IS1088 IS30 1103 3 6 ISRme10 IS30 1063 1 ISRme8 IS4 1455 1 1 ISRme5 IS481 1041 3 1 ISRme1 IS5 (IS427) 1331 2 2 ISRme6 IS5 (IS427) 913 1 ISRme7 IS6 840 2 ISRme19 IS66 2227 1 IS1071 Tn3 3204 3 1 * ISRme18 Tn3 ND 1 CHR chromosome. ND not determined * Copy inactivated by Tn6049 adjacent genes (Bentley et al. 2008;Bickhart et al. 2009;Hubner and Hendrickson 1997;Lin et al. 2007). Therefore, they are seen as a significant force in the adaptive and evolutionary response of their host, conferring genome plasticity that allows rapid adaptation to new environments (Mira et al. 2002;Schneider and Lenski 2004). ...
Cupriavidus metallidurans CH34 has been studied for over 30 years, mostly because of its resistance to numerous heavy metals. Many of these metal resistance determinants were rapidly associated with native megaplasmids. However, its genome sequencing and whole genome expression profiling not only revealed the complex structure of its multiple replicons and complex responses to metals, but also revealed the presence of unnoticed/unstudied metal resistance determinants on the different replicons. In this chapter, the genomic context of the metal response genes in C. metallidurans CH34 will be described with a focus on its mobilome including insertion sequence elements, transposons, integrative and conjugative elements and genomic islands.
... It has been shown that ISAba825 generated a 7-bp duplication (ATCGTTA) at the insertion site within carO [8]. It is well known that insertion sequences can cause mutations and genome rearrangements to enhance the spread of resistance genes and virulence determinants within the pathogenic species [10][11][12][13]. A recent report by Ravasi et al. [14] has shown the association and the tight regulation of ISAba825 with the plasmid-borne bla OXA-58 gene. ...
Clin Microbiol Infect 2012; 18: E446–E451
Four non-repetitive, clonally related (ST114), carbapenem-resistant Acinetobacter baumannii strains isolated in the USA were examined to understand the mechanisms of carbapenem resistance including screening for the presence of an insertion sequence upstream of the blaOXA-51-like gene, which could be involved in the control and expression of the antibiotic-resistance gene. We observed that the main mechanisms of carbapenem resistance were the result of the over-expression of the blaOXA-58-like and the blaOXA-65 gene, both of which had the presence of ISAba825 upstream of the genes. The importance of this element was shown by isolating plasmid-cured isogenic strains that had lost the plasmid with the ISAba825-blaOXA-58-like genes but during that same process also lost the chromosomal ISAba825 element present upstream of the blaOXA-65 gene. A 16-fold decrease in minimum inhibitory concentration of imipenem and an eight-fold decrease in the minimum inhibitory concentration of meropenem were seen in the isogenic strains that lost the plasmid. The study presents the first report of ISAba825 simultaneously governing the blaOXA-65 gene and the blaOXA-58-like gene expression and also highlights the importance of this element in carbapenem-sensitive isogenic strains, which were once carbapenem resistant.
... IS elements may also promote adaptation of their host genomes as demonstrated in experimental evolution experiments12131415. In addition, IS elements can influence or activate the expression of adjacent genes, e.g. by forming hybrid or fusion promoters or by containing outward-directed promoters16171819202122. IS elements are usually less than 2.5 kbp in length and have a relatively simple genetic organization. ...
... genes1718192021 , we investigated whether contiguous transcripts between A. asiaticus IS elements and downstream genes occur. We analyzed ten selected loci where IS elements and their downstream genes are encoded on the same strand and have the same orientation (Figure 4 ). ...
Insertion sequence (IS) elements are important mediators of genome plasticity and are widespread among bacterial and archaeal genomes. The 1.88 Mbp genome of the obligate intracellular amoeba symbiont Amoebophilus asiaticus contains an unusually large number of transposase genes (n = 354; 23% of all genes).
The transposase genes in the A. asiaticus genome can be assigned to 16 different IS elements termed ISCaa1 to ISCaa16, which are represented by 2 to 24 full-length copies, respectively. Despite this high IS element load, the A. asiaticus genome displays a GC skew pattern typical for most bacterial genomes, indicating that no major rearrangements have occurred recently. Additionally, the high sequence divergence of some IS elements, the high number of truncated IS element copies (n = 143), as well as the absence of direct repeats in most IS elements suggest that the IS elements of A. asiaticus are transpositionally inactive. Although we could show transcription of 13 IS elements, we did not find experimental evidence for transpositional activity, corroborating our results from sequence analyses. However, we detected contiguous transcripts between IS elements and their downstream genes at nine loci in the A. asiaticus genome, indicating that some IS elements influence the transcription of downstream genes, some of which might be important for host cell interaction.
Taken together, the IS elements in the A. asiaticus genome are currently in the process of degradation and largely represent reflections of the evolutionary past of A. asiaticus in which its genome was shaped by their activity.
... ISAba825 generated a 7-bp duplication (ATCGTTA) at the insertion site within carO (18). It is well known that ISs can cause insertion mutations and genome rearrangements and enhance the spread of resistance and virulence determinants within pathogenic species (2,9,11,13,17,19,23). In this work, we evaluated the impact of ISAba825 in modulating A. baumannii genome plasticity and carbapenem resistance. ...
ISAba825, an insertion sequence found inactivating Acinetobacter baumannii carO, was tagged with a kanamycin (Kn) resistance cassette. ISAba825::Kn effectively transposed in A. baumannii, showing preference for short, AT-enriched target sequences, generating 6- to 9-bp target duplications. Additionally, we
detected the presence of ISAba825 upstream of a plasmid-borne blaOXA-58 gene, generating a hybrid promoter largely enhancing its expression and leading to carbapenem resistance. Overall, a role
for ISAba825 in carbapenem resistance modulation in A. baumannii is proposed.
... IS ele- 94 ments have been implicated in the evolution of the host 95 as they contribute to diverse genomic rearrangements 96 (Bentley et al., 2008; Bickhart et al., 2009). Furthermore, 97 transposition could lead to altered gene expression, which 98 could be advantageous for the survival or the expression of 99 newly acquired genetic traits under certain conditions 100 (Hubner and Hendrickson, 1997; Lin et al., 2007). IS ele- 101 ments are therefore seen as a significant force in the adap- 102 tive and evolutionary response of their host, conferring 103 genome plasticity that allows rapid adaptation to new 104 environments (Mira et al., 2002; Schneider and Lenski, 105 2004). ...
Cupriavidus metallidurans CH34 is a β-proteobacterium well equipped to cope with harsh environmental conditions such as heavy metal pollution. The strain carries two megaplasmids specialized in the response to heavy metals and a considerable number of genomic islands, transposons and insertion sequence (IS) elements. The latter were characterized in detail in this study, which revealed nine new IS elements totaling to 21 distinct IS elements from 10 different IS families and reaching a total of 57 intact IS copies in CH34. Analysis of all fully sequenced bacterial genomes revealed that relatives of these IS elements were mostly found in the Burkholderiaceae family (β-proteobacteria) to which C. metallidurans belongs. Three IS elements were 100% conserved in other bacteria suggesting recent interaction and horizontal transfer between these strains. In addition, a number of these IS elements were associated with genomic islands, gene inactivation or rearrangements that alter the autotrophic growth capacities of CH34. The latter rearrangements gave the first molecular evidence for the mutator phenotype that is characteristic for various C. metallidurans strains. Furthermore, differential expression of some IS elements (or adjacent genes in the same strand orientation) was found under heavy metal stress, an environmental stress to which C. metallidurans CH34 is well adapted. These observations indicate that these IS elements play an active role in C. metallidurans CH34 lifestyle, including its metabolic potential and adaptation under selective pressure.
... Insertion sequences (IS) are the smallest and the most abundant transposable elements (Ͻ2.5 kb) capable of independent transposition in microbial genomes (20). They cause insertion mutations and genome rearrangements and enhance the spread of resistance and virulence determinants within species (2,12,16,18,23). Based on previous in silico analyses, ISAba1 is bracketed by 15-bp short inverted repeat sequences. ...
ISAba1 is an insertion sequence that is widely distributed in Acinetobacter baumannii. We demonstrated here that ISAba1 and the composite transposon Tn2006 are capable of transposition, generating 9-bp target site duplications. The expression of the ISAba1 transposase-encoding gene was downregulated by translational frameshifting.
... cynodontis (Lxc) is an endophytic bacterium originally isolated from Bermuda grass [11], which may serve as an engineered bacterium to improve the crop yield and quality [12]. Interestingly, during the course of studying the insertion sequence of Lxc, we have previously identified a truncated TA cassette fused with a partial segment of IS 1237 in pCXC100 [13]. ...
... Results 3.1. Cloning and sequence analysis of the talA/B cassette The complete copy of the talA/B cassette from the chromosome of the plasmid-free Lxc strain #3 was obtained by Tail- PCR using D26, D27, D28, D29 primers and AD1, AD2, AD3, AD4 arbitrary degenerate primers [13] ...
Here we report a toxin-antitoxin (TA) operon talAB identified from the Gram-positive bacterium Leifsonia xyli subsp. cynodontis. It is shown that talB encodes a broad-host cytotoxin functioning in different Gram-positive bacteria, while talA encodes its antidote. TalA and TalB form different hetero-oligomers in vitro; these hetero-oligomers, but not the antitoxin TalA, strongly bind to the talAB promoter region containing two inverted repeats. This represents a new mechanism of binding the promoter of a TA operon by the toxin and antitoxin complexes.
... Previously, 50 copies of five distinct IS elements (ISLxx1–5) in the Lxx genome were reported, along with 47 other transposase-related genes from uncharacterized elements [2,23]. In Lxc, three IS elements have previously been identified: IS1237 [24], ISLxc1 and ISLxc2 [25], located both on the chromosome and on the cryptic plasmid pCXC100 that is present in some isolates. A detailed characterization of two new Lxc IS elements (ISLxc3 and ISLxc4) and one new Lxx element (ISLxx6) is presented, as well as a comparative review of all elements found in both genomes, their distribution among families [11], and their comparative localization. ...
... IS1237 was first identified in the plasmid pCXC100 detected in some Lxc isolates [24]. Recently, 13 upstream and 10 downstream flanking sequences of IS1237 were described [25] . We identified 26 copies of IS1237 (Additional files 3 and 4) represented by two variant forms: 24 copies were 899-bp long and almost identical, with a single polymorphism in the 3'-IR; two copies were 798 bp long (Table 1 andFig. ...
... Since the coding sequence of all 899-bp long copies sequenced so far were identical, it is reasonable to assume that such translational features could be involved in the negative control of transposition [21]. The putative transposase has the expected catalytic domain containing a DDE motif [25], but the number of amino acids separating these residues does not follow the pattern of the IS5 family, and the glutamic acid is located downstream of the premature stop codon. Analyzing Lxx genome sequence, we identified ISLxx6, a single copy of which is represented within the genomic island LxxGI3 [2] . ...
Leifsonia xyli is a xylem-inhabiting bacterial species comprised of two subspecies: L. xyli subsp. xyli (Lxx) and L. xyli subsp. cynodontis (Lxc). Lxx is the causal agent of ratoon stunting disease in sugarcane commercial fields and Lxc colonizes the xylem of several grasses causing either mild or no symptoms of disease. The completely sequenced genome of Lxx provided insights into its biology and pathogenicity. Since IS elements are largely reported as an important source of bacterial genome diversification and nothing is known about their role in chromosome architecture of L. xyli, a comparative analysis of Lxc and Lxx elements was performed.
Sample sequencing of Lxc genome and comparative analysis with Lxx complete DNA sequence revealed a variable number of IS transposable elements acting upon genomic diversity. A detailed characterization of Lxc IS elements and a comparative review with IS elements of Lxx are presented. Each genome showed a unique set of elements although related to same IS families when considering features such as similarity among transposases, inverted and direct repeats, and element size. Most of the Lxc and Lxx IS families assigned were reported to maintain transposition at low levels using translation regulatory mechanisms, consistent with our in silico analysis. Some of the IS elements were found associated with rearrangements and specific regions of each genome. Differences were also found in the effect of IS elements upon insertion, although none of the elements were preferentially associated with gene disruption. A survey of transposases among genomes of Actinobacteria showed no correlation between phylogenetic relatedness and distribution of IS families. By using Southern hybridization, we suggested that diversification of Lxc isolates is also mediated by insertion sequences in probably recent events.
Collectively our data indicate that transposable elements are involved in genome diversification of Lxc and Lxx. The IS elements were probably acquired after the divergence of the two subspecies and are associated with genome organization and gene contents. In addition to enhancing understanding of IS element dynamics in general, these data will contribute to our ongoing comparative analyses aimed at understanding the biological differences of the Lxc and Lxx.
