Evaluation of effectiveness of bacteriophage purification methods
Abstract The use of bacteriophages for therapy has increased over the last decade. While there is need for clear regulatory pathways for bacteriophage approval for mainstream use in clinical practice, practitioners and patients have been able to access bacteriophage therapy under compassionate groun...
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| Language: | English |
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BMC
2024-12-01
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| Series: | Virology Journal |
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| Online Access: | https://doi.org/10.1186/s12985-024-02580-y |
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| author | Siti Saleha Binte Mohamed Yakob Adil Joseph Tucci Helen Irving Cassandra Cianciarulo Mwila Kabwe |
| author_facet | Siti Saleha Binte Mohamed Yakob Adil Joseph Tucci Helen Irving Cassandra Cianciarulo Mwila Kabwe |
| author_sort | Siti Saleha Binte Mohamed Yakob Adil |
| collection | DOAJ |
| description | Abstract The use of bacteriophages for therapy has increased over the last decade. While there is need for clear regulatory pathways for bacteriophage approval for mainstream use in clinical practice, practitioners and patients have been able to access bacteriophage therapy under compassionate grounds and through magistral preparations. However, there is currently no standard for purifying these bacteriophages to ensure safety, and good manufacturing practice certification may not be achieved in these emergency uses. In this study, we employed an Interleukin Receptor Associated Kinase (IRAK) 3 knockout monocyte-based assay to evaluate the endotoxin removal efficacy of three common bacteriophage purification methods: Triton X-100 exposure, CsCl density gradient ultracentrifugation, and Pierce™ High-Capacity Endotoxin Removal Resin spin columns. In our experiments we tested these purification methods on three different bacteriophage morphotypes: siphovirus, podovirus and myovirus. We showed that the lowest endotoxin levels and immune responses were achieved when purifying bacteriophages with Triton-X treatment. The results from purifying with CsCl density gradient ultracentrifugation were comparable, and these were both significantly better than purification with Pierce™ High-Capacity Endotoxin Removal Resin spin columns. We also showed that Triton X-100 purification resulted in the lowest loss of bacteriophage titres. Finally, of the bacteriophages tested here, it did not appear that virus morphology affected efficacy of endotoxin removal. |
| format | Article |
| id | doaj-art-dee698b2781949eabf245799b0b7a153 |
| institution | Kabale University |
| issn | 1743-422X |
| language | English |
| publishDate | 2024-12-01 |
| publisher | BMC |
| record_format | Article |
| series | Virology Journal |
| spelling | doaj-art-dee698b2781949eabf245799b0b7a1532024-12-22T12:15:01ZengBMCVirology Journal1743-422X2024-12-0121111010.1186/s12985-024-02580-yEvaluation of effectiveness of bacteriophage purification methodsSiti Saleha Binte Mohamed Yakob Adil0Joseph Tucci1Helen Irving2Cassandra Cianciarulo3Mwila Kabwe4Department of Rural Clinical Sciences, La Trobe Rural Health School, La Trobe UniversityDepartment of Rural Clinical Sciences, La Trobe Rural Health School, La Trobe UniversityDepartment of Rural Clinical Sciences, La Trobe Rural Health School, La Trobe UniversityDepartment of Rural Clinical Sciences, La Trobe Rural Health School, La Trobe UniversityDepartment of Rural Clinical Sciences, La Trobe Rural Health School, La Trobe UniversityAbstract The use of bacteriophages for therapy has increased over the last decade. While there is need for clear regulatory pathways for bacteriophage approval for mainstream use in clinical practice, practitioners and patients have been able to access bacteriophage therapy under compassionate grounds and through magistral preparations. However, there is currently no standard for purifying these bacteriophages to ensure safety, and good manufacturing practice certification may not be achieved in these emergency uses. In this study, we employed an Interleukin Receptor Associated Kinase (IRAK) 3 knockout monocyte-based assay to evaluate the endotoxin removal efficacy of three common bacteriophage purification methods: Triton X-100 exposure, CsCl density gradient ultracentrifugation, and Pierce™ High-Capacity Endotoxin Removal Resin spin columns. In our experiments we tested these purification methods on three different bacteriophage morphotypes: siphovirus, podovirus and myovirus. We showed that the lowest endotoxin levels and immune responses were achieved when purifying bacteriophages with Triton-X treatment. The results from purifying with CsCl density gradient ultracentrifugation were comparable, and these were both significantly better than purification with Pierce™ High-Capacity Endotoxin Removal Resin spin columns. We also showed that Triton X-100 purification resulted in the lowest loss of bacteriophage titres. Finally, of the bacteriophages tested here, it did not appear that virus morphology affected efficacy of endotoxin removal.https://doi.org/10.1186/s12985-024-02580-yBacteriophageEndotoxin purificationTriton X-100Caesium chlorideEndotoxin removal resin |
| spellingShingle | Siti Saleha Binte Mohamed Yakob Adil Joseph Tucci Helen Irving Cassandra Cianciarulo Mwila Kabwe Evaluation of effectiveness of bacteriophage purification methods Virology Journal Bacteriophage Endotoxin purification Triton X-100 Caesium chloride Endotoxin removal resin |
| title | Evaluation of effectiveness of bacteriophage purification methods |
| title_full | Evaluation of effectiveness of bacteriophage purification methods |
| title_fullStr | Evaluation of effectiveness of bacteriophage purification methods |
| title_full_unstemmed | Evaluation of effectiveness of bacteriophage purification methods |
| title_short | Evaluation of effectiveness of bacteriophage purification methods |
| title_sort | evaluation of effectiveness of bacteriophage purification methods |
| topic | Bacteriophage Endotoxin purification Triton X-100 Caesium chloride Endotoxin removal resin |
| url | https://doi.org/10.1186/s12985-024-02580-y |
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