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Performance of a 70-mer oligonucleotide microarray for genotyping of Campylobacter jejuni.

Rodin S, Andersson AF, Wirta V, Eriksson L, Ljungström M, Björkholm B, Lindmark H, Engstrand L - BMC Microbiol. (2008)

Bottom Line: We further identify regions that frequently differ between isolates, including both previously described and novel regions.Finally, we show that genes that belong to certain functional groups differ between isolates more often than expected by chance.In this study we demonstrated the utility of 70-mer oligonucleotide microarrays for genotyping of Campylobacter jejuni isolates, with resolution outperforming MLST.

View Article: PubMed Central - HTML - PubMed

Affiliation: Swedish Institute for Infectious Disease Control, SE-17182 Solna, Sweden. sandra.rodin@smi.ki.se

ABSTRACT

Background: Campylobacter jejuni is widespread in the environment and is the major cause of bacterial gastroenteritis in humans. In the present study we use microarray-based comparative genomic hybridizations (CGH), pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST) to analyze closely related C. jejuni isolates from chicken and human infection.

Results: With the exception of one isolate, the microarray data clusters the isolates according to the five groups determined by PFGE. In contrast, MLST defines only three genotypes among the isolates, indicating a lower resolution. All methods show that there is no inherit difference between isolates infecting humans and chicken, suggesting a common underlying population of C. jejuni. We further identify regions that frequently differ between isolates, including both previously described and novel regions. Finally, we show that genes that belong to certain functional groups differ between isolates more often than expected by chance.

Conclusion: In this study we demonstrated the utility of 70-mer oligonucleotide microarrays for genotyping of Campylobacter jejuni isolates, with resolution outperforming MLST.

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Related in: MedlinePlus

Analysis of variable regions in the C. jejuni genome. (A) Hierarchical clustering (Euclidian distance, average linking) based on the entire CGH dataset. The probes are ordered according to their position in the genome, starting with the probe targeting gene Cj0001 at the bottom of the figure. The log2 ratio (M-values) of each probe is represented using a color gradient from yellow to blue, denoting M-values ranging from 0 to -7. A negative M-value indicates that the probe shows sequence divergence or absence in the test isolate compared to the NCTC 11168 reference strain. The PFGE genotypes and MLST sequence types are shown below. (B) Presence of variable regions in the analyzed isolates. On the y-axis are the five types of isolates (A, B, C, D and H) included in the study. The height of the bars corresponds to the number of isolate types in which the variable region was identified. The width of each bar is drawn proportional to the number of genes in the corresponding variable region. Regions 1–16 were described by Taboada et al. [27] and regions 17–18 by Parker et al. [29]. Regions 19–21 were identified in this study. (C) Analysis of the divergent genes shows that these represent multiple COG groups, as exemplified for isolate C20. Description of the COG groups is available through the COG database [33]. (D) The representation of COG groups among the variable genes was analyzed using Fisher exact test and the results are summarized using color coding. Groups M and V were significantly overrepresented among the variable genes in multiple isolates.
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Figure 4: Analysis of variable regions in the C. jejuni genome. (A) Hierarchical clustering (Euclidian distance, average linking) based on the entire CGH dataset. The probes are ordered according to their position in the genome, starting with the probe targeting gene Cj0001 at the bottom of the figure. The log2 ratio (M-values) of each probe is represented using a color gradient from yellow to blue, denoting M-values ranging from 0 to -7. A negative M-value indicates that the probe shows sequence divergence or absence in the test isolate compared to the NCTC 11168 reference strain. The PFGE genotypes and MLST sequence types are shown below. (B) Presence of variable regions in the analyzed isolates. On the y-axis are the five types of isolates (A, B, C, D and H) included in the study. The height of the bars corresponds to the number of isolate types in which the variable region was identified. The width of each bar is drawn proportional to the number of genes in the corresponding variable region. Regions 1–16 were described by Taboada et al. [27] and regions 17–18 by Parker et al. [29]. Regions 19–21 were identified in this study. (C) Analysis of the divergent genes shows that these represent multiple COG groups, as exemplified for isolate C20. Description of the COG groups is available through the COG database [33]. (D) The representation of COG groups among the variable genes was analyzed using Fisher exact test and the results are summarized using color coding. Groups M and V were significantly overrepresented among the variable genes in multiple isolates.

Mentions: We next carried out a hierarchical clustering analysis using the microarray data to identify similarities among the isolates. The origin of the isolates (chicken or human) had no effect on the clustering. Instead, the isolates clustered into groups similar to those obtained by PFGE and MLST. Three major clusters were identified (Figure 4A). The first included all isolates with PFGE types A, B and C, belonging to the MLST ST-21 clonal complex. Within this cluster, the two isolates of PFGE type B clustered together, while another cluster was formed by the two type C and three type A isolates. The remaining type A isolate clustered outside this tight cluster. The microarray data indicated that this cluster of isolates is similar to the reference strain NCTC 11168 (Figure 4A). This relatedness is further supported by the MLST data; strain NCTC 11168 belongs to the same clonal complex as the type A, B and C isolates (ST-21 clonal complex), although it is of a different sequence type (ST-43). The second cluster included PFGE type D isolates (MLST clonal complex ST-677) and strain RM1221, and the third PFGE type H isolates (MLST clonal complex ST-48).


Performance of a 70-mer oligonucleotide microarray for genotyping of Campylobacter jejuni.

Rodin S, Andersson AF, Wirta V, Eriksson L, Ljungström M, Björkholm B, Lindmark H, Engstrand L - BMC Microbiol. (2008)

Analysis of variable regions in the C. jejuni genome. (A) Hierarchical clustering (Euclidian distance, average linking) based on the entire CGH dataset. The probes are ordered according to their position in the genome, starting with the probe targeting gene Cj0001 at the bottom of the figure. The log2 ratio (M-values) of each probe is represented using a color gradient from yellow to blue, denoting M-values ranging from 0 to -7. A negative M-value indicates that the probe shows sequence divergence or absence in the test isolate compared to the NCTC 11168 reference strain. The PFGE genotypes and MLST sequence types are shown below. (B) Presence of variable regions in the analyzed isolates. On the y-axis are the five types of isolates (A, B, C, D and H) included in the study. The height of the bars corresponds to the number of isolate types in which the variable region was identified. The width of each bar is drawn proportional to the number of genes in the corresponding variable region. Regions 1–16 were described by Taboada et al. [27] and regions 17–18 by Parker et al. [29]. Regions 19–21 were identified in this study. (C) Analysis of the divergent genes shows that these represent multiple COG groups, as exemplified for isolate C20. Description of the COG groups is available through the COG database [33]. (D) The representation of COG groups among the variable genes was analyzed using Fisher exact test and the results are summarized using color coding. Groups M and V were significantly overrepresented among the variable genes in multiple isolates.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2396164&req=5

Figure 4: Analysis of variable regions in the C. jejuni genome. (A) Hierarchical clustering (Euclidian distance, average linking) based on the entire CGH dataset. The probes are ordered according to their position in the genome, starting with the probe targeting gene Cj0001 at the bottom of the figure. The log2 ratio (M-values) of each probe is represented using a color gradient from yellow to blue, denoting M-values ranging from 0 to -7. A negative M-value indicates that the probe shows sequence divergence or absence in the test isolate compared to the NCTC 11168 reference strain. The PFGE genotypes and MLST sequence types are shown below. (B) Presence of variable regions in the analyzed isolates. On the y-axis are the five types of isolates (A, B, C, D and H) included in the study. The height of the bars corresponds to the number of isolate types in which the variable region was identified. The width of each bar is drawn proportional to the number of genes in the corresponding variable region. Regions 1–16 were described by Taboada et al. [27] and regions 17–18 by Parker et al. [29]. Regions 19–21 were identified in this study. (C) Analysis of the divergent genes shows that these represent multiple COG groups, as exemplified for isolate C20. Description of the COG groups is available through the COG database [33]. (D) The representation of COG groups among the variable genes was analyzed using Fisher exact test and the results are summarized using color coding. Groups M and V were significantly overrepresented among the variable genes in multiple isolates.
Mentions: We next carried out a hierarchical clustering analysis using the microarray data to identify similarities among the isolates. The origin of the isolates (chicken or human) had no effect on the clustering. Instead, the isolates clustered into groups similar to those obtained by PFGE and MLST. Three major clusters were identified (Figure 4A). The first included all isolates with PFGE types A, B and C, belonging to the MLST ST-21 clonal complex. Within this cluster, the two isolates of PFGE type B clustered together, while another cluster was formed by the two type C and three type A isolates. The remaining type A isolate clustered outside this tight cluster. The microarray data indicated that this cluster of isolates is similar to the reference strain NCTC 11168 (Figure 4A). This relatedness is further supported by the MLST data; strain NCTC 11168 belongs to the same clonal complex as the type A, B and C isolates (ST-21 clonal complex), although it is of a different sequence type (ST-43). The second cluster included PFGE type D isolates (MLST clonal complex ST-677) and strain RM1221, and the third PFGE type H isolates (MLST clonal complex ST-48).

Bottom Line: We further identify regions that frequently differ between isolates, including both previously described and novel regions.Finally, we show that genes that belong to certain functional groups differ between isolates more often than expected by chance.In this study we demonstrated the utility of 70-mer oligonucleotide microarrays for genotyping of Campylobacter jejuni isolates, with resolution outperforming MLST.

View Article: PubMed Central - HTML - PubMed

Affiliation: Swedish Institute for Infectious Disease Control, SE-17182 Solna, Sweden. sandra.rodin@smi.ki.se

ABSTRACT

Background: Campylobacter jejuni is widespread in the environment and is the major cause of bacterial gastroenteritis in humans. In the present study we use microarray-based comparative genomic hybridizations (CGH), pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST) to analyze closely related C. jejuni isolates from chicken and human infection.

Results: With the exception of one isolate, the microarray data clusters the isolates according to the five groups determined by PFGE. In contrast, MLST defines only three genotypes among the isolates, indicating a lower resolution. All methods show that there is no inherit difference between isolates infecting humans and chicken, suggesting a common underlying population of C. jejuni. We further identify regions that frequently differ between isolates, including both previously described and novel regions. Finally, we show that genes that belong to certain functional groups differ between isolates more often than expected by chance.

Conclusion: In this study we demonstrated the utility of 70-mer oligonucleotide microarrays for genotyping of Campylobacter jejuni isolates, with resolution outperforming MLST.

Show MeSH
Related in: MedlinePlus