A Genomic Island of the Pathogen Leptospira interrogans Serovar Lai Can Excise from Its Chromosome

Abstract

An examination of the two Leptospira interrogans genomes sequenced so far reveals few genetic differences, including an extra DNA region, 54 kb in length, in L. interrogans serovar Lai. This locus contains 103 predicted coding sequences that are absent from the genome of L. interrogans serovar Copenhageni, of which only 20% had significant BLASTP hits in GenBank. By analyzing the L. interrogans serovar Lai genome by pulsed-field gel electrophoresis, we also found that this 54-kb DNA fragment exists as a circular plasmid. This was confirmed by amplification of a DNA fragment corresponding to that of the predicted fragment if this region excised from the chromosome and its left and right ends joined together. In addition, cloning of the putative rep gene of this DNA region was responsible for autonomous replication in Leptospira spp., therefore generating a new Escherichia coli-Leptospira sp. shuttle vector. Taken together, our results show that this genomic island can excise from the chromosome and form a replicative plasmid. Analysis of the distribution of this genomic island revealed that highly related sequences exist in other L. interrogans virulent strains. This genomic island, containing a high proportion of novel genes, may have an important role in spreading genes, including virulence factors, among bacterial populations.

Spontaneous Excision of the Salmonella enterica Serovar Enteritidis-Specific Defective Prophage-Like Element φSE14

“…Excision of GEIs from the bacterial chromosome often leads to the formation of a detectable extrachromosomal CF (6,9,23,34,43). Although inverse PCR and sequence analyses indicate that spontaneous excision of SE14 most likely results in the formation of a CF, we do not yet have direct physical evidence to prove the existence of such an excision product, as in the case of other unstable GEIs (4,6).…”

“…Although inverse PCR and sequence analyses indicate that spontaneous excision of SE14 most likely results in the formation of a CF, we do not yet have direct physical evidence to prove the existence of such an excision product, as in the case of other unstable GEIs (4,6). Furthermore, in addition to the possibility of CF formation, we cannot yet exclude the possibility that SE14 can also exist as tandem repeats in the bacterial chromosome, as our inverse PCR products are also consistent with this possibility.…”

Comparative proteogenomic analysis of the Leptospira interrogans virulence-attenuated strain IPAV against the pathogenic strain 56601

“…Second, the sequence of an approximately 6.5-kb DNA region spanning the revised CI coordinates 3 248 961 to 3 255 503 was erroneously reversed in the originally published Lai 56601 genome map. Four other large fragments were rectified, including sequence rectification from 414 967 to 415 363 (LA_0417), a sequence addition from 2 051 058 to 2 051 577 (LA_2081a and LA_2083) with respect to the revised CI coordinates, a sequence addition from 332 934 to 333 358 (LB_340 and LB_340a) with respect to the revised CII coordinates, and a deletion of an 831-bp direct repeat flanking the Genome Island I (GI I) [16]. Finally, 43 single nucleotides were rectified according to the resequencing of the corresponding sites.…”

A Replicative Plasmid Vector Allows Efficient Complementation of Pathogenic Leptospira Strains

“…Markers for the selection of transformants include kanamycin, spectinomycin, and gentamicin resistance cassettes (3,5,6). The replication origins of L. biflexa phage LE1 (3), L. biflexa plasmid p74 (7), and a phage-related genomic island from L. interrogans (8) have previously been used to construct L. biflexa-E. coli plasmid shuttle vectors. However, no shuttle vector construct for intermediate or pathogenic Leptospira spp.…”