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International audience; Oenococcus oeni is responsible for the malolactic fermentation of wines. Genomic diversity has been recently established in the species and extensive attention is now being given to the genomic bases of strain-specific differences. We explored the role of insertion sequences (IS), which are considered as driving forces for novel genotypic and phenotypic variants in prokaryotes. The present study focuses on members of the IS30 family, which are widespread among lactic acid bacteria. An in silico analysis of the three available genomes of O. oeni in combination with the use of an inverse PCR strategy targeting conserved IS30-related sequences indicated the presence of seven IS30 copies in the pangenome of O. oeni. A primer designed to anneal to the conserved 3' end of the IS30 element was paired with each of the seven primers selected to bind to unique sequences upstream of each of the seven mobile elements identified. The study presents an overview of the abundance, and the genomic environment of IS30 elements in the O. oeni pangenome and shows that the two existing genetic sub-populations previously described in the species through multilocus sequence typing analysis (MLST) differ in their IS30 content. Possible IS30 impacts on bacterial adaptation are discussed.

Summary Objectives The aim of the present study was to assess the usefulness of insertion element IS 1395 for differentiation of Mycobacterium xenopi , an increasingly common opportunistic human pathogen. Methods Fifty-two isolates obtained from 51 patients in Poland in 1996, 1997, and 1999, were analyzed by IS 1395 restriction fragment length polymorphism (RFLP), and their susceptibilities to 11 anti-tuberculosis drugs were also determined. Results IS 1395 -associated banding patterns of the isolates were not highly polymorphic; the RFLP patterns displayed several bands in common. Nevertheless, 44 of the 52 isolates were clearly distinguishable from each other. Only eight strains (15.4%) occurred in four clusters of two identical clones, one of which comprised two isolates obtained from one patient with a 12-month interval. The remaining six patients with clustered strains showed no apparent epidemiologic links with the other patients from the same cluster, and they were most likely infected by the same environmental source. No noticeable difference in RFLP pattern or IS 1395 copy number between drug-sensitive and drug-resistant strains was shown. A high proportion (84.6%) of strains resistant to at least one drug was found, and 7.7% were resistant to more than three drugs. Conclusions The stability and utility of IS 1395 for further detailed epidemiological investigations of M. xenopi was confirmed and extended.

We developed a novel PCR-fingerprinting system for differentiation of enterobacterial strains using a single oligonucleotide primer IS1tr that matches the inverted terminal repeats of the IS1 insertion element. Compared to widely used BOX-PCR and ribotyping methods, our system features higher resolution allowing differentiation of closely related isolates that appear identical in BOX-PCR and ribotyping but differ in their phage sensitivity. The IS1-profiling system is less sensitive to the quality of the material and equipment used. At the same time, BOX-PCR is more universal and suitable for bacterial strain grouping and reconstruction of the low-distance phylogeny. Thus, our system represents an important supplement to the existing set of tools for bacterial strain differentiation; it is particularly valuable for a detailed investigation of highly divergent and rapidly evolving natural bacterial populations and for studies on coliphage ecology. However, some isolates could not be reliably differentiated by IS1-PCR, because of the low number of bands in their patterns. For improvement of IS1-fingerprinting characteristics, we offer to modify the system by introducing the second primer TR8834 hybridizing to the sequence of a transposase gene that is widely spread in enterobacterial genomes.

The first sul2 genes have been found in S. maltophilia from several different countries., Trimethoprim/sulfamethoxazole (TMP/SMX) resistance remains a serious threat in the treatment of Stenotrophomonas maltophilia infections. We analyzed an international collection of 55 S. maltophilia TMP/SMX-sensitive (S) (n = 30) and -resistant (R) (n = 25) strains for integrons; sul1, sul2 and dhfr genes; and insertion element common region (ISCR) elements. sul1, as part of a class 1 integron, was detected in 17 of 25 TMP/SMX-R. Nine TMP/SMX-R strains carried sul2; 7 were on large plasmids. Five TMP/SMX-R isolates were positive for ISCR2, and 4 were linked to sul2; 2 others possessed ISCR3. Two ISCR2s were adjacent to floR. Six TMP/SMX-S isolates harbored novel ISCR elements, ISCR9 and ISCR10. Linkage of ISCR3, ISCR9, and ISCR10 to sul2 and dhfr genes was not demonstrated. The data from this study indicate that class 1 integrons and ISCR elements linked to sul2 genes can mediate TMP/SMX resistance in S. maltophilia and are geographically widespread, findings that reinforce the need for ongoing resistance surveillance.

Transposon Tn 2610 , found in a conjugative plasmid from an Escherichia coli isolate recovered at a hospital in Chiba, Japan, in 1975, was completely sequenced. Tn 2610 is 23,883 bp long and is bracketed by two transposition modules, a Tn 1721 -like module and a Tn 21 -derived module, which correspond, respectively, to the long inverted repeats IRa and IRb previously described for this transposon. Although both tnpA genes are intact, only that in the Tn 21 -derived module (IRb) functions in the transposition, while that in the Tn 1721 -derived module (IRa) cannot recognize the 38-bp imperfect repeat at the end of the IRb element. Both tnpR and res are present in IRa, while the tnpR gene of IRb is interrupted by the insertion of an IS 26 insertion element. The intervening region, between the res site of the Tn 1721 module and IS 26 , carries multiple integron-associated resistance genes within a Tn 21 backbone, including a region identical to that found in the genome of Salmonella enterica serovar Typhimurium DT104. These findings suggest that Tn 2610 originated from Tn 1721 and Tn 21 , with extensive recombination events with other elements which have resulted in a complex mosaic structure.

An IS element, termed ISCg2, was identified in the chromosome of Corynebacterium glutamicum ATCC 13032. After screening a cosmid library of the C. glutamicum ATCC 13032 genome, six copies of ISCg2 including their flanking regions were sequenced and analyzed. ISCg2 is 1636 bp in length and has 26-bp imperfect inverted repeats flanked by 3-bp direct repeats. By comparisons with other IS elements,ISCg2 was classified as a member of the IS30 family of insertion sequences. The six copies of ISCg2 were identical at the nucleotide level and were located in intergenic, AT-rich regions of the chromosome. The regions in which the six copies of ISCg2 were inserted displayed significant similarities. This similarity extends over a region of 65 bp, which was assumed to be the target region for ISCg2. Interestingly, live of the six copies of ISCg2 were located adjacent to genes that may be involved in aspartate and glutamate metabolism or its regulation. Investigation of the distribution of ISCg2 showed that the IS element is restricted to certain C. glutamicum strains. Analysis of various integration regions indicates active transposition of ISCg2. in C. glutamicum.

The blood samples of 66 Ahmadiyya Muslim males from Qadian, district Gurdaspur of Punjab have been analysed to study the Y-chromosome Alu insertion polymorphism (YAP). Y-chromosomes carrying the YAP element were found in many populations in India and Pakistan. However, the absence of YAP insertion element in the present Ahmadiyya population has suggested that this isolated population is not likely to have been affected by many migrations in Indian history.

The SWI/SNF-like chromatin-remodeling protein ATRX has emerged as a key factor in the regulation of α-globin gene expression, incorporation of histone variants into the chromatin template and, more recently, as a frequently mutated gene across a wide spectrum of cancers. Therefore, the availability of a functional ATRX cDNA for expression studies is a valuable tool for the scientific community. We have identified two independent transposon insertions of a bacterial IS10 element into exon 8 of ATRX isoform 2 coding sequence in two different plasmids derived from a single source. We demonstrate that these insertion events are common and there is an insertion hotspot within the ATRX cDNA. Such IS10 insertions produce a truncated form of ATRX, which significantly compromises its nuclear localization. In turn, we describe ways to prevent IS10 insertion during propagation and cloning of ATRX-containing vectors, including optimal growth conditions, bacterial strains, and suggested sequencing strategies. Finally, we have generated an insertion-free plasmid that is available to the community for expression studies of ATRX. Electronic supplementary material The online version of this article (doi:10.1186/2193-1801-3-222) contains supplementary material, which is available to authorized users.

Comparing strains within the same microbial species has proven effective in the identification of genes and genomic regions responsible for virulence, as well as in the diagnosis and treatment of infectious diseases. In this paper, we present Sibelia, a tool for finding synteny blocks in multiple closely related microbial genomes using iterative de Bruijn graphs. Unlike most other tools, Sibelia can find synteny blocks that are repeated within genomes as well as blocks shared by multiple genomes. It represents synteny blocks in a hierarchy structure with multiple layers, each of which representing a different granularity level. Sibelia has been designed to work efficiently with a large number of microbial genomes; it finds synteny blocks in 31 S. aureus genomes within 31 minutes and in 59 E.coli genomes within 107 minutes on a standard desktop. Sibelia software is distributed under the GNU GPL v2 license and is available at: https://github.com/bioinf/Sibelia. Sibelia’s web-server is available at: http://etool.me/software/sibelia.

The nucleotide sequence of an insertion element in R-factor R1767 derivative pBP11 was determined. It is almost overall identical with IS15 delta, IS26 and IS46. Like IS46 it flanks one end of the sul-bla determinant and is involved in amplification of the resistance cassette. The significance for this process of a palindrome comprising part of IS15 delta IV is discussed.

The species Campylobacter fetus is divided into the subspecies C. fetus subsp. venerealis (CFV) and C. fetus subsp. fetus (CFF). CFV is the causative agent of bovine genital campylobacteriosis, a highly contagious venereal disease that may lead to serious reproductive problems, including sterility and abortion. In contrast, CFF can be isolated from the gastrointestinal tract of a wide range of host species, is associated with abortion in sheep and cattle, and can also be isolated from local and systemic infections in humans. Despite differences in host and niche preferences, microbiological differentiation of the two subspecies of C. fetus is extremely difficult. This study describes the identification of a new insertion element, ISCfe1, which is present exclusively in CFV strains, with highly conserved specific ISCfe1 insertion sites. The results are useful for identification and differentiation of the two C. fetus subspecies and will help in understanding the evolution and pathogenesis of C. fetus.

International audience; Some strains of Lactobacillus sakei are known to produce the bacteriocin sakacin P, encoded by the spp gene cluster. In strains unable to produce sakacin P, spp homologues were observed. The analysis of 15 strains not producing sakacin P revealed that all contained a region corresponding to a part of sppKR encoding the regulatory elements for sakacin P production. In some strains homologues of sppE and sppT, responsible for sakacin P transport, and the sakacin P structural gene sppA and its immunity gene spiA, were also present. The sequence of the chromosomal spp-related gene cluster was determined in two non-producing strains: L. sakei Lb790 and L. sakei 23K. The L. sakei Lb790 spp gene cluster encompasses genes homologous to sppK, sppR, sppT and sppE. In L. sakei 23K, only sppK and sppR homologues were present. The sppK homologues appeared non-functional as they contained mutations and/or an insertion element. In addition to the spp homologues, several small putative genes were found in the gene clusters of the two strains. Some were similar in both strains, and their organization suggests a mosaic structure resulting from successive rearrangements. Transcriptional analysis showed that the genes of the L. sakei Lb790 spp cluster were expressed when genes encoding an operative sakacin P regulatory system were introduced in this strain, thus complementing the inactive sppK gene. Expression experiments also suggested that some of the spp homologues maintained their function in non-producing strains.

These examples of observations of sequences leave more questions than answers but provide intriguing hints that the metabolism of this important group of bacteria is versatile and complex.

The Institut Pasteur in Paris, located at the very same premises since it was founded by its namesake in 1888, was originally meant to produce immunizing sera and consequently developed a long and sound tradition in medical bacteriology and virology. Through the latter, the institute was a venue for phage research of the pre-Delbruck era. It was Felix d’Herelle, a French-Canadian researcher at the Institut Pasteur, who described what he called in 1917 bacteriophages which he isolated from the stools of patients convalescing from shigellosis disentery. Accordingly, d’Herelle is considered to be one of the discoverers of phages. Although the British Frederick Twort had, already in 1915, observed a bacteria-killing, self-reproducing agent in culture plates with staphyloccoci, this was not known to d’Herelle. Expert opinions diverge on to whom the merit of priority should go. While Twort’s work ended abruptly with his death as a soldier during the First World War, d’Herelle left a row of followers who proceeded to describe a series of different phenomena related to phages, but never succeeded in really interpreting their observations. Years later, the husband-and-wife team of Eugene and Elisabeth Wollman, also working at the Institut Pasteur, noticed a remarkable phenomenon: under certain circumstances, some phages would infect bacteria without causing their disintegration. Such phages faded from immediate detection, but each cell from the resulting clone stayed under the constant threat, albeit with low probability, of lysing after some generations, liberating progeny phages in the process. This was the phenomenon called lysogeny. Aside from the virulent phages which caused, unconditionally, the infected bacteria to lyse, there existed, apparently, another category of phages called temperate, which were able to enter the host, and then establish and maintain with it a state of nonlethal equilibrium for many generations. These viral entities — nowadays called provirus or prophages — divided synchronously with the host for many cell cycles, retaining nevertheless the capability of initiating a full-blown lytic cycle. Lysogenic bacteria, i.e. those harboring a prophage, live under the Damoclean sword of the all-time imminent, although rarely spontaneously occurring, death by lysis.

A new insertion sequence (IS2112) was identified in the genome of the 1-haloalkane-utilizing bacterium Rhodococcus rhodochrous NCIMB 13064. The insertion element is 1415 bp long, does not contain terminal inverted repeats, and is not flanked by directly repeated sequences. IS2112 belongs to the IS110 family of transposable elements, and forms a separate subfamily, along with IS116, Two copies of IS2112 were found in R, rhodochrous NCIMB 13064 and one, two or three copies of a similar sequence were detected in five other 1-haloalkane-degrading Rhodococcus strains. There were no sequences homologous to IS2112 found in the l-haloalkane-degrading 'Pseudomonas pavonaceae' 170 and Rhodococcus sp, HA1 or in several Rhodococcus strains which do not utilize haloalkanes, IS2112 was originally found in plasmid pRTL1 of R. rhodochrous NCIMB 13064 which harbours genes encoding utilization of l-haloalkanes, and was located 5 kbp upstream of the haloalkane dehalogenase gene (dhaA), Although the second copy of IS2112 in strain NCIMB 13064 was also present on the pRTL1 plasmid, these sequences do not apparently comprise a single composite transposon encoding haloalkane utilization. An analysis of derivatives of NCIMB 13064 revealed that IS2112 was involved in genome rearrangements. IS2112 appeared to change its location as a result of transposition and as a result of other rearrangements of the NCIMB 13064 genome.

The effects of two mutagens ENU and MMS characterized by different alkylation patterns have been studied on the reversion of an MR-induced singed mutation to wild-type. Reversion of this unstable singed mutation under the influence of MR is assumed to represent the removal or transposition of an insertion element. Since MR acts primarily in spermatogonia, the mutagens were fed to 1st instar larvae. Recessive lethal tests were carried out simultaneously to calibrate for the mutagenic effectiveness of the chemicals. For both powerful mutagens, it was observed that the frequency of reversion remained far below of what would have been expected on the basis of the mutagenic effectiveness, as registered in the lethal tests. Thus 1 mM ENU, 5 mM and 10 mM MMS did not affect the reversion frequency at all, and with 3 mM ENU only a doubling of the reversion frequency was observed, despite a 5-fold increase in the lethal frequency. The threshold at 1 mM EMU and the low effectiveness of 3 mM on the reversion process are taken as an indication that ENU affected the transposition process in an indirect manner, rather than the excision events themselves. The data obtained with Drosophila are consistent with the microbial observations in that mutation involving removal or transposition of an insertion element is not affected by mutagenic treatments. This finding may have consequences for the evaluation of induced genetic damage on the basis of the spontaneous load of genetic detriment in man. An incidental observation was that non-MR Cy larvae exhibited greater sensitivity to the induction of recessive lethals by MMS than MR-individuals.

The nucleotide sequence of the insertion element IS200 has been determined partially, including the junctions between the element and the host chromosome at the insertion site. At most, two bases (A-A) are found repeated at the junctions and could be duplications of host sequences generated by the insertion of the element. No obvious sequence repeats, either direct or inverted, have been detected between the sequences just within the two ends of the element. The element is an extremely strong block to host transcription across the insertion site. A sequence similar to known transcription termination signals was found just within the element near the right end. Removal of less than 50 base-pairs at the right end of the element abolishes the transcription block. The putative terminator sequence is located within this 50 base-pair region. Genetic studies suggest that the element contains a promoter located more than 93 base-pairs from its left end. The proposed promoter and terminator are in proper orientation to form an internal transcription unit.

Recently a new insertion element (IS102)b ha been described in plasmid pSC101. We have determined its complete sequence: it consists of 1057 bp; 338 bp at one end are identical to those already determined for the kanamycin resistance transposon Tn903. It is not flanked by any direct repeat. Its coding capabilities are discussed, and compared to those of IS903.

Genomic clones from a Northern Flint Line of Zea mays are shown to carry a 700 bp insertion, Cin 1. The Cin 1 insert is not present in the homologous, unique DNA fragment of a midwestern maize line. However, Cin 1 is present in multiple copies in both lines.

Promoters located within the Tn10 insertion element cause transcription of "host" sequences adjacent to both ends of the inserted Tn10 element. These promoters are usually not observed in genetic experiments because their transcripts are efficiently terminated at nearby rho-dependent termination sites. The observations presented here provide an explanation for several confusing aspects of transposon behavior and suggest the possibility that many transposons possess promoters that have escaped detection for similar reasons.

MR is a frequently occurring mutator in Drosophila melanogaster inducing mutation by the incorporation of insertion sequences. In the presence of MR a mutation at the singed (sn) locus induced by MR, reverts to wild-type at a high frequency of 1.7%. This reversion system which presumably involves the removal of an insertion element, was used to study the effects of defective DNA repair. Thus, reversion frequencies were compared in progeny of flies with mei-9, deficient for excision repair, mei-41, deficient for post-replication repair, or with both mei-9 and mei-41. The data show that under conditions of defective DNA repair, the frequency of MR-mediated reversion, is consistently decreased in comparison to repair-proficient conditions. This effect is explained by assuming that defective repair interferes with some steps in the process of reverse mutation involving the removal of insertion sequences. The observed reduction in reversion frequency may well result from selective elimination of cells in which the reversion process has not been completed.

Southern blotting using a 5′-proximal probe of the Saccharomyces cerevisiae CYR1 gene has revealed heterogeneity in laboratory strains. It is demonstrated that strain AB320 contains a Ty1 element inserted in the promoter region of CYR1. The Ty1 orientation suggests that transcription of CYR1 is initiated downstream from the insertion region.

Two previously characterized duplications in the insertion element IS2 of Escherichia coli can be described as end-products resulting from endonucleolytic cleavages of DNA at regions of potential secondary structure and subsequent DNA synthesis.

IS1-mediated insertion and deletion formation occur preferentially into A+T-rich regions of DNA of bacteriophate P1 and of the r-determinant of the R plasmid NR1. The significance of this correlation is discussed in view of other published data.

Total DNAs from twelve natural isolates of Escherichia coli from animals and humans were examined by hybridization with a probe for IS1. Considerable variation in copy number was found. In the case of two strains isolated from the same individual, one strain contained no copies of IS1 and the other, much greater than 30. Evidence was also obtained for the existence of IS1-like elements (iso-IS1s) of greater than 15% sequence divergence relative to the IS1 from antibiotic resistance plasmid R100.

A comparison of the available lambdoid origin sequences allows to point out several structural features in common with the DNA sequence of the phage G4 intercistronic region between genes F and G, which can be interpreted as recognition elements of a primase promoter sequence. These leading strand initiation signals are located in leftward orientation on the A-rich strand of all lambdoid origins, and by comparison with G4 require a single-stranded binding sequence, which appears to result from transcriptional activation. Binding of 0 protein to the origin interferes with transcription across ori , which therefore, has to precede these ori/0 interactions.

The study of transposable elements and the highly mutable alleles they create allows recognition of the fluid nature of both prokaryotic and eukaryotic genomes. Mutations at particular alleles have been instrumental in dissecting the spatial and temporal patterns of gene expression in maize (Zea mays L.) and Drosophila. And with the advent of molecular cloning, the mutable alleles containing a particular insertion element can be isolated by virtue of the transposon “tag” they contain. Recently we have begun studying the genetic and molecular properties of mutable alleles derived from maize lines containing Mutator activity. In this paper we review the properties of unstable alleles of maize and the genetic properties of Mutator lines of maize. A new transposable element family has been discovered and isolated from an unstable allele recovered from a Mutator line (Barker, et al., 1984). The molecular properties of this element called Mu-l will be reviewed.

The insertion element ISR1 was isolated from the soil bacterium R. lupini. In this strain, ISR1 shows a very strong affinity to plasmid RP4. It causes RP4 mutations at the strikingly high frequency of 10(-2) to 10(-1), either by the integration itself or by generating deletions. In E. coli, ISR1 seems to be inactive. No evidence could be obtained for a promoter site on ISR1 or for an ISR1-encoded protein. Our results indicate, however, an ISR1-specific termination signal for either transcription or translation.

The insertion sequence, IS1, is among the smallest insertion elements known. It is a member of a class of elements which includes IS10, IS50, and IS903 which are frequently found in pairs flanking a nontransposable DNA segment. In concert, the 2 flanking elements are able to promote transposition of the entire structure. This type of transposable element has been called a compound or composite transposon. The best characterized IS1-flanked compound transposon is perhaps Tn9 in which 2 directly repeated copies of IS1 flank a 1.1-kb segment of DNA which encodes chloramphenicol acetyltransferase (CAT) (1). Unlike most other elements of this class, however, IS1 is present in the chromosomes of a variety of enterobacteria, in some cases in many copies (2).

Four different strains of Escherichia coli and several other bacteria were examined by Southern analysis for the presence of the insertion element IS5 and IS5-like sequences. Variations in the copy number, degree of homology and restriction pattern of the IS5-like sequence were found among the different organisms. The number and distribution of IS5 sequences do not appear to correlate with the evolutionary relationship of the bacteria in which they occur.