Improvement of the Pour Plate Method by Separate Sterilization of Agar and Other Medium Components and Reduction of the Agar Concentration
ABSTRACT Although the pour plate method is widely employed in microbiological quality control, it has certain drawbacks, including having to melt the culture medium before seeding. In this study, the preparation of the culture medium was modified by using a lower concentration of agar (10 g/L), whic...
| Main Authors: | , , , , , , , , , , , |
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| Format: | Article |
| Language: | English |
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American Society for Microbiology
2023-02-01
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| Series: | Microbiology Spectrum |
| Subjects: | |
| Online Access: | https://journals.asm.org/doi/10.1128/spectrum.03161-22 |
| _version_ | 1811164395492868096 |
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| author | I. Terrones-Fernandez P. Casino A. López S. Peiró S. Ríos A. Nardi-Ricart E. García-Montoya D. Asensio A. M. Marqués R. Castilla P. J. Gamez-Montero N. Piqué |
| author_facet | I. Terrones-Fernandez P. Casino A. López S. Peiró S. Ríos A. Nardi-Ricart E. García-Montoya D. Asensio A. M. Marqués R. Castilla P. J. Gamez-Montero N. Piqué |
| author_sort | I. Terrones-Fernandez |
| collection | DOAJ |
| description | ABSTRACT Although the pour plate method is widely employed in microbiological quality control, it has certain drawbacks, including having to melt the culture medium before seeding. In this study, the preparation of the culture medium was modified by using a lower concentration of agar (10 g/L), which was separated from the nutrients during sterilization. The new protocol was assessed in media frequently used in microbiological quality control of food, cosmetics, and pharmaceutical products, with tryptic soy agar (TSA), Sabouraud 4% dextrose agar (SDA), and violet red bile glucose agar (VRBG). In comparison with the conventionally produced media, the modifications significantly improved the growth of Saccharomyces cerevisiae in SDA, Staphylococcus aureus, Salmonella enterica subsp. enterica serovar Typhimurium, and Candida albicans in TSA and Escherichia coli ATCC 8739 and ATCC 25922 and S. Typhimurium in VRBG. The modified VRBG was also more selective for Pseudomonas aeruginosa. Regarding physicochemical properties, a significantly lower pH was observed in TSA and VRBG and lower strength values in TSA. Sterilizing agar separately from the other components of the medium and reducing the agar concentration to 10 g/L can improve microorganism growth and enhance the selectivity of differential media in the pour plate method. These modifications could facilitate the automation of this culture technique. IMPORTANCE In the era of rapid microbiological methods, there is a need to improve long-established culture techniques. Drawbacks of the pour plate method include having to melt each medium separately before seeding. For this technique, we demonstrate that separating the agar from the other components of commonly used media during sterilization and reducing the agar concentration to 10 g/L can enhance microbial growth. The new protocol could have advantages in routine laboratory practice because less agar is required and the same molten agar suspension can be used to prepare different media. Moreover, these modifications could facilitate the automation of the pour plate method. |
| first_indexed | 2024-04-10T15:20:50Z |
| format | Article |
| id | doaj.art-bfbba950d8fc49caa515ccda7b07c041 |
| institution | Directory Open Access Journal |
| issn | 2165-0497 |
| language | English |
| last_indexed | 2024-04-10T15:20:50Z |
| publishDate | 2023-02-01 |
| publisher | American Society for Microbiology |
| record_format | Article |
| series | Microbiology Spectrum |
| spelling | doaj.art-bfbba950d8fc49caa515ccda7b07c0412023-02-14T14:15:50ZengAmerican Society for MicrobiologyMicrobiology Spectrum2165-04972023-02-0111110.1128/spectrum.03161-22Improvement of the Pour Plate Method by Separate Sterilization of Agar and Other Medium Components and Reduction of the Agar ConcentrationI. Terrones-Fernandez0P. Casino1A. López2S. Peiró3S. Ríos4A. Nardi-Ricart5E. García-Montoya6D. Asensio7A. M. Marqués8R. Castilla9P. J. Gamez-Montero10N. Piqué11Department of Quality Control, Reactivos para Diagnóstico, S.L. (RPD), Barcelona, Catalonia, SpainDepartment of Quality Control, Reactivos para Diagnóstico, S.L. (RPD), Barcelona, Catalonia, SpainDepartment of Quality Control, Reactivos para Diagnóstico, S.L. (RPD), Barcelona, Catalonia, SpainDepartment of Quality Control, Reactivos para Diagnóstico, S.L. (RPD), Barcelona, Catalonia, SpainDepartment of Genetics, Microbiology and Statistics, Biology Faculty, Universitat de Barcelona, Barcelona, Catalonia, SpainDepartment of Pharmacy and Pharmaceutical Technology and Physical Chemistry, Faculty of Pharmacy and Food Sciences, Universitat de Barcelona, Catalonia, SpainDepartment of Pharmacy and Pharmaceutical Technology and Physical Chemistry, Faculty of Pharmacy and Food Sciences, Universitat de Barcelona, Catalonia, SpainDepartment of Quality Control, Reactivos para Diagnóstico, S.L. (RPD), Barcelona, Catalonia, SpainMicrobiology Section, Department of Biology, Healthcare and Environment, Faculty of Pharmacy and Food Sciences, Universitat de Barcelona (UB), Barcelona, Catalonia, SpainCATMech. Department of Fluid Mechanics, Universitat Politecnica de Catalunya, Terrassa, Catalonia, SpainCATMech. Department of Fluid Mechanics, Universitat Politecnica de Catalunya, Terrassa, Catalonia, SpainMicrobiology Section, Department of Biology, Healthcare and Environment, Faculty of Pharmacy and Food Sciences, Universitat de Barcelona (UB), Barcelona, Catalonia, SpainABSTRACT Although the pour plate method is widely employed in microbiological quality control, it has certain drawbacks, including having to melt the culture medium before seeding. In this study, the preparation of the culture medium was modified by using a lower concentration of agar (10 g/L), which was separated from the nutrients during sterilization. The new protocol was assessed in media frequently used in microbiological quality control of food, cosmetics, and pharmaceutical products, with tryptic soy agar (TSA), Sabouraud 4% dextrose agar (SDA), and violet red bile glucose agar (VRBG). In comparison with the conventionally produced media, the modifications significantly improved the growth of Saccharomyces cerevisiae in SDA, Staphylococcus aureus, Salmonella enterica subsp. enterica serovar Typhimurium, and Candida albicans in TSA and Escherichia coli ATCC 8739 and ATCC 25922 and S. Typhimurium in VRBG. The modified VRBG was also more selective for Pseudomonas aeruginosa. Regarding physicochemical properties, a significantly lower pH was observed in TSA and VRBG and lower strength values in TSA. Sterilizing agar separately from the other components of the medium and reducing the agar concentration to 10 g/L can improve microorganism growth and enhance the selectivity of differential media in the pour plate method. These modifications could facilitate the automation of this culture technique. IMPORTANCE In the era of rapid microbiological methods, there is a need to improve long-established culture techniques. Drawbacks of the pour plate method include having to melt each medium separately before seeding. For this technique, we demonstrate that separating the agar from the other components of commonly used media during sterilization and reducing the agar concentration to 10 g/L can enhance microbial growth. The new protocol could have advantages in routine laboratory practice because less agar is required and the same molten agar suspension can be used to prepare different media. Moreover, these modifications could facilitate the automation of the pour plate method.https://journals.asm.org/doi/10.1128/spectrum.03161-22pour plate methodTSASDAVRBGrecoveryselectivity |
| spellingShingle | I. Terrones-Fernandez P. Casino A. López S. Peiró S. Ríos A. Nardi-Ricart E. García-Montoya D. Asensio A. M. Marqués R. Castilla P. J. Gamez-Montero N. Piqué Improvement of the Pour Plate Method by Separate Sterilization of Agar and Other Medium Components and Reduction of the Agar Concentration Microbiology Spectrum pour plate method TSA SDA VRBG recovery selectivity |
| title | Improvement of the Pour Plate Method by Separate Sterilization of Agar and Other Medium Components and Reduction of the Agar Concentration |
| title_full | Improvement of the Pour Plate Method by Separate Sterilization of Agar and Other Medium Components and Reduction of the Agar Concentration |
| title_fullStr | Improvement of the Pour Plate Method by Separate Sterilization of Agar and Other Medium Components and Reduction of the Agar Concentration |
| title_full_unstemmed | Improvement of the Pour Plate Method by Separate Sterilization of Agar and Other Medium Components and Reduction of the Agar Concentration |
| title_short | Improvement of the Pour Plate Method by Separate Sterilization of Agar and Other Medium Components and Reduction of the Agar Concentration |
| title_sort | improvement of the pour plate method by separate sterilization of agar and other medium components and reduction of the agar concentration |
| topic | pour plate method TSA SDA VRBG recovery selectivity |
| url | https://journals.asm.org/doi/10.1128/spectrum.03161-22 |
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