Examination of clinical and environmental Vibrio parahaemolyticus isolates by multi-locus sequence typing (MLST) and multiple-locus variable-number tandem-repeat analysis (MLVA)
Vibrio parahaemolyticus is a leading cause of seafood-borne infections in the US. This organism has a high genetic diversity that complicates identification of strain relatedness and epidemiological investigations. However, sequence-based analysis methods are promising tools for these identificati...
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| Format: | Article |
| Language: | English |
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Frontiers Media S.A.
2015-06-01
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| Series: | Frontiers in Microbiology |
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| Online Access: | http://journal.frontiersin.org/Journal/10.3389/fmicb.2015.00564/full |
| _version_ | 1818502288363225088 |
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| author | Catharina H.M. Lüdeke Catharina H.M. Lüdeke Narjol eGonzález-Escalona Markus eFischer Jessica L Jones |
| author_facet | Catharina H.M. Lüdeke Catharina H.M. Lüdeke Narjol eGonzález-Escalona Markus eFischer Jessica L Jones |
| author_sort | Catharina H.M. Lüdeke |
| collection | DOAJ |
| description | Vibrio parahaemolyticus is a leading cause of seafood-borne infections in the US. This organism has a high genetic diversity that complicates identification of strain relatedness and epidemiological investigations. However, sequence-based analysis methods are promising tools for these identifications. In this study, Multi-Locus Sequence Typing (MLST) and Multiple-Locus Variable-Number Tandem-Repeat Analysis (MLVA) was performed on 58 V. parahaemolyticus isolates (28 of oyster and 30 of clinical origin), to identify differences in phylogeny. The results obtained by both methods were compared to Pulsed-Field Gel Electrophoresis (PFGE) patterns determined in a previous study. Forty-one unique sequence types (STs) were identified by MLST among the 58 isolates. Almost half of the isolates (22) belonged to a new ST and added to the MLST database. A ST could not be generated for 5 (8.6%) isolates, primarily due to an untypable recA locus. Analysis with eBURST did not identify any clonal complex among the strains analyzed and revealed 37 singeltons with 4 of them forming 2 groups (1 of them SLV, and the other a DLV). An established MLVA assay, targeting 12 total genes through three separate 4-plex PCRs, was successfully adapted to high resolution melt (HRM) analysis with faster and easier experimental setup; resulting in 58 unique melt curve patterns. HRM-MLVA was capable of differentiating isolates within the same PFGE cluster and having the same ST. Conclusively, combining the three methods PFGE, MLST, and HRM-MLVA, for the phylogenetic analysis of V. parahaemolyticus resulted in a high resolution subtyping scheme for V. parahaemolyticus. This scheme will be useful as a phylogenetic research tool and as an improved method for outbreak investigations for V. parahaemolyticus. |
| first_indexed | 2024-12-10T21:08:15Z |
| format | Article |
| id | doaj.art-6cd1d54bbba346dc93117633242ca203 |
| institution | Directory Open Access Journal |
| issn | 1664-302X |
| language | English |
| last_indexed | 2024-12-10T21:08:15Z |
| publishDate | 2015-06-01 |
| publisher | Frontiers Media S.A. |
| record_format | Article |
| series | Frontiers in Microbiology |
| spelling | doaj.art-6cd1d54bbba346dc93117633242ca2032022-12-22T01:33:33ZengFrontiers Media S.A.Frontiers in Microbiology1664-302X2015-06-01610.3389/fmicb.2015.00564145591Examination of clinical and environmental Vibrio parahaemolyticus isolates by multi-locus sequence typing (MLST) and multiple-locus variable-number tandem-repeat analysis (MLVA)Catharina H.M. Lüdeke0Catharina H.M. Lüdeke1Narjol eGonzález-Escalona2Markus eFischer3Jessica L Jones4United Stated Food and Drug AdministrationUniversity of HamburgUnited States Food and Drug AdministrationUniversity of HamburgUnited Stated Food and Drug AdministrationVibrio parahaemolyticus is a leading cause of seafood-borne infections in the US. This organism has a high genetic diversity that complicates identification of strain relatedness and epidemiological investigations. However, sequence-based analysis methods are promising tools for these identifications. In this study, Multi-Locus Sequence Typing (MLST) and Multiple-Locus Variable-Number Tandem-Repeat Analysis (MLVA) was performed on 58 V. parahaemolyticus isolates (28 of oyster and 30 of clinical origin), to identify differences in phylogeny. The results obtained by both methods were compared to Pulsed-Field Gel Electrophoresis (PFGE) patterns determined in a previous study. Forty-one unique sequence types (STs) were identified by MLST among the 58 isolates. Almost half of the isolates (22) belonged to a new ST and added to the MLST database. A ST could not be generated for 5 (8.6%) isolates, primarily due to an untypable recA locus. Analysis with eBURST did not identify any clonal complex among the strains analyzed and revealed 37 singeltons with 4 of them forming 2 groups (1 of them SLV, and the other a DLV). An established MLVA assay, targeting 12 total genes through three separate 4-plex PCRs, was successfully adapted to high resolution melt (HRM) analysis with faster and easier experimental setup; resulting in 58 unique melt curve patterns. HRM-MLVA was capable of differentiating isolates within the same PFGE cluster and having the same ST. Conclusively, combining the three methods PFGE, MLST, and HRM-MLVA, for the phylogenetic analysis of V. parahaemolyticus resulted in a high resolution subtyping scheme for V. parahaemolyticus. This scheme will be useful as a phylogenetic research tool and as an improved method for outbreak investigations for V. parahaemolyticus.http://journal.frontiersin.org/Journal/10.3389/fmicb.2015.00564/fullVibrio parahaemolyticusphylogeneticsmlstMLVAHRM |
| spellingShingle | Catharina H.M. Lüdeke Catharina H.M. Lüdeke Narjol eGonzález-Escalona Markus eFischer Jessica L Jones Examination of clinical and environmental Vibrio parahaemolyticus isolates by multi-locus sequence typing (MLST) and multiple-locus variable-number tandem-repeat analysis (MLVA) Frontiers in Microbiology Vibrio parahaemolyticus phylogenetics mlst MLVA HRM |
| title | Examination of clinical and environmental Vibrio parahaemolyticus isolates by multi-locus sequence typing (MLST) and multiple-locus variable-number tandem-repeat analysis (MLVA) |
| title_full | Examination of clinical and environmental Vibrio parahaemolyticus isolates by multi-locus sequence typing (MLST) and multiple-locus variable-number tandem-repeat analysis (MLVA) |
| title_fullStr | Examination of clinical and environmental Vibrio parahaemolyticus isolates by multi-locus sequence typing (MLST) and multiple-locus variable-number tandem-repeat analysis (MLVA) |
| title_full_unstemmed | Examination of clinical and environmental Vibrio parahaemolyticus isolates by multi-locus sequence typing (MLST) and multiple-locus variable-number tandem-repeat analysis (MLVA) |
| title_short | Examination of clinical and environmental Vibrio parahaemolyticus isolates by multi-locus sequence typing (MLST) and multiple-locus variable-number tandem-repeat analysis (MLVA) |
| title_sort | examination of clinical and environmental vibrio parahaemolyticus isolates by multi locus sequence typing mlst and multiple locus variable number tandem repeat analysis mlva |
| topic | Vibrio parahaemolyticus phylogenetics mlst MLVA HRM |
| url | http://journal.frontiersin.org/Journal/10.3389/fmicb.2015.00564/full |
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