Limits...
Conservation of the S10-spc-alpha locus within otherwise highly plastic genomes provides phylogenetic insight into the genus Leptospira.

Victoria B, Ahmed A, Zuerner RL, Ahmed N, Bulach DM, Quinteiro J, Hartskeerl RA - PLoS ONE (2008)

Bottom Line: In contrast, PCR analysis of this locus using DNA from saprophytic Leptospira species and species with an intermediate pathogenic capacity generated varied results.Multilocus sequence typing (MLST) of four conserved regions resulted in the construction of well-defined phylogenetic trees that help resolve questions about the interrelationships of pathogenic Leptospira.Based on the results of secY sequence analysis, we found that reliable species identification of pathogenic Leptospira is possible by comparative analysis of a 245 bp region commonly used as a target for diagnostic PCR for leptospirosis.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Molecular Biology, Faculty of Biology, University of Santiago de Compostela, Galicia, Spain.

ABSTRACT
S10-spc-alpha is a 17.5 kb cluster of 32 genes encoding ribosomal proteins. This locus has an unusual composition and organization in Leptospira interrogans. We demonstrate the highly conserved nature of this region among diverse Leptospira and show its utility as a phylogenetically informative region. Comparative analyses were performed by PCR using primer sets covering the whole locus. Correctly sized fragments were obtained by PCR from all L. interrogans strains tested for each primer set indicating that this locus is well conserved in this species. Few differences were detected in amplification profiles between different pathogenic species, indicating that the S10-spc-alpha locus is conserved among pathogenic Leptospira. In contrast, PCR analysis of this locus using DNA from saprophytic Leptospira species and species with an intermediate pathogenic capacity generated varied results. Sequence alignment of the S10-spc-alpha locus from two pathogenic species, L. interrogans and L. borgpetersenii, with the corresponding locus from the saprophyte L. biflexa serovar Patoc showed that genetic organization of this locus is well conserved within Leptospira. Multilocus sequence typing (MLST) of four conserved regions resulted in the construction of well-defined phylogenetic trees that help resolve questions about the interrelationships of pathogenic Leptospira. Based on the results of secY sequence analysis, we found that reliable species identification of pathogenic Leptospira is possible by comparative analysis of a 245 bp region commonly used as a target for diagnostic PCR for leptospirosis. Comparative analysis of Leptospira strains revealed that strain H6 previously classified as L. inadai actually belongs to the pathogenic species L. interrogans and that L. meyeri strain ICF phylogenetically co-localized with the pathogenic clusters. These findings demonstrate that the S10-spc-alpha locus is highly conserved throughout the genus and may be more useful in comparing evolution of the genus than loci studied previously.

Show MeSH

Related in: MedlinePlus

Phylogenetic trees based on Tamura-Nei distances and elaborated using the Neighbor-Joining method.Distances were calculated from the 300–301 (A), 621–625 (B), 624–650 (C) and G1–G2 (D) sequence fragments within the S10-spc-α locus of pathogenic species of Leptospira. The total evidence was combined and analyzed under identical conditions (E). In addition, data available from 16S rDNA (rrs) sequences were used to obtain an alternative hypothesis for the relationships of diverse Leptospira strains (F). Dotted lines show alternative branching patterns, with bootstrapping values ≥50%, obtained in the consensus majority rule tree obtained by parsimony criterion. Numbers above branches represent the percentage of bootstrapping results (2000 replicates). Trees are drawn to scale as indicated by the bar depicted below each tree; bars represent the estimated distance in units of the number of base substitutions per site. The scale the 16S rRNA-based tree is expanded relative to other loci. L. biflexa was used as the outgroup.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2481283&req=5

pone-0002752-g002: Phylogenetic trees based on Tamura-Nei distances and elaborated using the Neighbor-Joining method.Distances were calculated from the 300–301 (A), 621–625 (B), 624–650 (C) and G1–G2 (D) sequence fragments within the S10-spc-α locus of pathogenic species of Leptospira. The total evidence was combined and analyzed under identical conditions (E). In addition, data available from 16S rDNA (rrs) sequences were used to obtain an alternative hypothesis for the relationships of diverse Leptospira strains (F). Dotted lines show alternative branching patterns, with bootstrapping values ≥50%, obtained in the consensus majority rule tree obtained by parsimony criterion. Numbers above branches represent the percentage of bootstrapping results (2000 replicates). Trees are drawn to scale as indicated by the bar depicted below each tree; bars represent the estimated distance in units of the number of base substitutions per site. The scale the 16S rRNA-based tree is expanded relative to other loci. L. biflexa was used as the outgroup.

Mentions: In the composite tree (Fig. 2E), pathogenic strains were separated into two well-supported clades that are similar, but not identical to clades resolved in the binary tree. One clade consists of the sister sub-clades containing L. interrogans and L. noguchii, with L. kirschneri located in a basal position. This clade is consistently recovered in all topologies (Fig. 2), with the exception of the tree based on the 621–625 fragment (Fig. 2B) and parsimony topology generated from G1–G2 sequences (Fig. 2D), where L. kirschneri and L. noguchii swap their positions. The close relationships of these species are also apparent through comparative analysis using 16S rDNA sequence data (Fig. 2F) and independent binary data (Fig. 1). The second clade of the composite tree contains L. borgpetersenii, L. santarosai, L, alexanderi, L. weilii and L. meyeri strain ICF. Although, the branching pattern within this clade has lower support, the sibling relationship between L. alexanderi and L. weilii is well conserved. The relative positions of L. borgpetersenii, L. meyeri and L. santarosai are uncertain and vary depending on the data set and method of analysis (Fig. 2). In the tree inferred from the G1–G2 locus, the Celledoni and Sarmin strains of L. weilii are located in separate clades.


Conservation of the S10-spc-alpha locus within otherwise highly plastic genomes provides phylogenetic insight into the genus Leptospira.

Victoria B, Ahmed A, Zuerner RL, Ahmed N, Bulach DM, Quinteiro J, Hartskeerl RA - PLoS ONE (2008)

Phylogenetic trees based on Tamura-Nei distances and elaborated using the Neighbor-Joining method.Distances were calculated from the 300–301 (A), 621–625 (B), 624–650 (C) and G1–G2 (D) sequence fragments within the S10-spc-α locus of pathogenic species of Leptospira. The total evidence was combined and analyzed under identical conditions (E). In addition, data available from 16S rDNA (rrs) sequences were used to obtain an alternative hypothesis for the relationships of diverse Leptospira strains (F). Dotted lines show alternative branching patterns, with bootstrapping values ≥50%, obtained in the consensus majority rule tree obtained by parsimony criterion. Numbers above branches represent the percentage of bootstrapping results (2000 replicates). Trees are drawn to scale as indicated by the bar depicted below each tree; bars represent the estimated distance in units of the number of base substitutions per site. The scale the 16S rRNA-based tree is expanded relative to other loci. L. biflexa was used as the outgroup.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2481283&req=5

pone-0002752-g002: Phylogenetic trees based on Tamura-Nei distances and elaborated using the Neighbor-Joining method.Distances were calculated from the 300–301 (A), 621–625 (B), 624–650 (C) and G1–G2 (D) sequence fragments within the S10-spc-α locus of pathogenic species of Leptospira. The total evidence was combined and analyzed under identical conditions (E). In addition, data available from 16S rDNA (rrs) sequences were used to obtain an alternative hypothesis for the relationships of diverse Leptospira strains (F). Dotted lines show alternative branching patterns, with bootstrapping values ≥50%, obtained in the consensus majority rule tree obtained by parsimony criterion. Numbers above branches represent the percentage of bootstrapping results (2000 replicates). Trees are drawn to scale as indicated by the bar depicted below each tree; bars represent the estimated distance in units of the number of base substitutions per site. The scale the 16S rRNA-based tree is expanded relative to other loci. L. biflexa was used as the outgroup.
Mentions: In the composite tree (Fig. 2E), pathogenic strains were separated into two well-supported clades that are similar, but not identical to clades resolved in the binary tree. One clade consists of the sister sub-clades containing L. interrogans and L. noguchii, with L. kirschneri located in a basal position. This clade is consistently recovered in all topologies (Fig. 2), with the exception of the tree based on the 621–625 fragment (Fig. 2B) and parsimony topology generated from G1–G2 sequences (Fig. 2D), where L. kirschneri and L. noguchii swap their positions. The close relationships of these species are also apparent through comparative analysis using 16S rDNA sequence data (Fig. 2F) and independent binary data (Fig. 1). The second clade of the composite tree contains L. borgpetersenii, L. santarosai, L, alexanderi, L. weilii and L. meyeri strain ICF. Although, the branching pattern within this clade has lower support, the sibling relationship between L. alexanderi and L. weilii is well conserved. The relative positions of L. borgpetersenii, L. meyeri and L. santarosai are uncertain and vary depending on the data set and method of analysis (Fig. 2). In the tree inferred from the G1–G2 locus, the Celledoni and Sarmin strains of L. weilii are located in separate clades.

Bottom Line: In contrast, PCR analysis of this locus using DNA from saprophytic Leptospira species and species with an intermediate pathogenic capacity generated varied results.Multilocus sequence typing (MLST) of four conserved regions resulted in the construction of well-defined phylogenetic trees that help resolve questions about the interrelationships of pathogenic Leptospira.Based on the results of secY sequence analysis, we found that reliable species identification of pathogenic Leptospira is possible by comparative analysis of a 245 bp region commonly used as a target for diagnostic PCR for leptospirosis.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Molecular Biology, Faculty of Biology, University of Santiago de Compostela, Galicia, Spain.

ABSTRACT
S10-spc-alpha is a 17.5 kb cluster of 32 genes encoding ribosomal proteins. This locus has an unusual composition and organization in Leptospira interrogans. We demonstrate the highly conserved nature of this region among diverse Leptospira and show its utility as a phylogenetically informative region. Comparative analyses were performed by PCR using primer sets covering the whole locus. Correctly sized fragments were obtained by PCR from all L. interrogans strains tested for each primer set indicating that this locus is well conserved in this species. Few differences were detected in amplification profiles between different pathogenic species, indicating that the S10-spc-alpha locus is conserved among pathogenic Leptospira. In contrast, PCR analysis of this locus using DNA from saprophytic Leptospira species and species with an intermediate pathogenic capacity generated varied results. Sequence alignment of the S10-spc-alpha locus from two pathogenic species, L. interrogans and L. borgpetersenii, with the corresponding locus from the saprophyte L. biflexa serovar Patoc showed that genetic organization of this locus is well conserved within Leptospira. Multilocus sequence typing (MLST) of four conserved regions resulted in the construction of well-defined phylogenetic trees that help resolve questions about the interrelationships of pathogenic Leptospira. Based on the results of secY sequence analysis, we found that reliable species identification of pathogenic Leptospira is possible by comparative analysis of a 245 bp region commonly used as a target for diagnostic PCR for leptospirosis. Comparative analysis of Leptospira strains revealed that strain H6 previously classified as L. inadai actually belongs to the pathogenic species L. interrogans and that L. meyeri strain ICF phylogenetically co-localized with the pathogenic clusters. These findings demonstrate that the S10-spc-alpha locus is highly conserved throughout the genus and may be more useful in comparing evolution of the genus than loci studied previously.

Show MeSH
Related in: MedlinePlus