Construction and characterization of bioluminescent Salmonella Reading outbreak and non-outbreak strains
ABSTRACT Salmonella enterica serotype Reading has recently been identified as a significant foodborne pathogen from contaminated poultry products. There is a critical need for close monitoring of this newly emerged pathogen. This study developed bioluminescent strains of S. Reading for real-time pat...
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| Format: | Article |
| Language: | English |
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American Society for Microbiology
2025-02-01
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| Series: | Microbiology Spectrum |
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| Online Access: | https://journals.asm.org/doi/10.1128/spectrum.01263-24 |
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| author | Abubakar Shitu Isah Reshma Ramachandran Anuraj Theradiyil Sukumaran Aaron S. Kiess Claudia D. Castañeda Timothy Boltz Kenneth S. Macklin Hossam Abdelhamed Li Zhang |
| author_facet | Abubakar Shitu Isah Reshma Ramachandran Anuraj Theradiyil Sukumaran Aaron S. Kiess Claudia D. Castañeda Timothy Boltz Kenneth S. Macklin Hossam Abdelhamed Li Zhang |
| author_sort | Abubakar Shitu Isah |
| collection | DOAJ |
| description | ABSTRACT Salmonella enterica serotype Reading has recently been identified as a significant foodborne pathogen from contaminated poultry products. There is a critical need for close monitoring of this newly emerged pathogen. This study developed bioluminescent strains of S. Reading for real-time pathogen tracking using bioluminescence imaging. Two strains of S. Reading were used: an outbreak strain and a non-outbreak strain. The chloramphenicol acetyltransferase gene was cloned into the plasmid pBS-slpGFPluxABCDE, which carries luxABCDE operon and an ampicillin resistance gene. The newly constructed plasmid was then transformed into the outbreak and non-outbreak strains of S. Reading by electroporation. The resulting colonies were confirmed by visualizing bioluminescence using an in vivo imaging system and by testing their resistance to chloramphenicol. These strains demonstrated a high bioluminescence level (108–109 Photons/s/cm2/sr) and were tested for growth and plasmid stability by daily subculturing in Luria–Bertani medium with and without antibiotics. The plasmid remained stable for 8 days under non-selective conditions, and growth rates were comparable to non-bioluminescent parent strains in antibiotic-free conditions. However, growth was notably different in the presence of chloramphenicol, indicating successful plasmid retention and function. This study successfully created stable bioluminescent S. Reading strains, marking a significant step forward in monitoring and potentially reducing the spread of this emergent foodborne pathogen in the poultry industry.IMPORTANCESalmonella enterica serotype Reading has recently become a significant foodborne pathogen linked to poultry products. To enhance pathogen monitoring, this study developed bioluminescent strains of S. Reading by inserting the chloramphenicol acetyltransferase gene into a plasmid containing a bioluminescence gene cluster. These modified strains were transformed into outbreak and non-outbreak bacterial strains via electroporation. The bioluminescent strains demonstrated stable plasmid retention and high bioluminescence levels. They also showed growth comparable to their parent strains, even in the absence of antibiotics. These bioluminescent strains could potentially facilitate real-time monitoring and control of S. Reading in poultry industries. |
| format | Article |
| id | doaj-art-225afcdef55342aeb557135392d12cfc |
| institution | Kabale University |
| issn | 2165-0497 |
| language | English |
| publishDate | 2025-02-01 |
| publisher | American Society for Microbiology |
| record_format | Article |
| series | Microbiology Spectrum |
| spelling | doaj-art-225afcdef55342aeb557135392d12cfc2025-02-04T14:03:41ZengAmerican Society for MicrobiologyMicrobiology Spectrum2165-04972025-02-0113210.1128/spectrum.01263-24Construction and characterization of bioluminescent Salmonella Reading outbreak and non-outbreak strainsAbubakar Shitu Isah0Reshma Ramachandran1Anuraj Theradiyil Sukumaran2Aaron S. Kiess3Claudia D. Castañeda4Timothy Boltz5Kenneth S. Macklin6Hossam Abdelhamed7Li Zhang8Department of Poultry Science, Mississippi State University, Mississippi State, Mississippi, USAIDEXX Laboratories, One IDEXX Drive, Westbrook, Maine, USAFreshpet, Bethlehem, Pennsylvania, USAPrestage Department of Poultry Science, North Carolina State University, Raleigh, North Carolina, USAEngrain LLC, Manhattan, Kansas, USADepartment of Poultry Science, Mississippi State University, Mississippi State, Mississippi, USADepartment of Poultry Science, Mississippi State University, Mississippi State, Mississippi, USADepartment of Comparative Biomedical Sciences, College Veterinary Medicine, Mississippi State University, Mississippi State, Mississippi, USADepartment of Poultry Science, Mississippi State University, Mississippi State, Mississippi, USAABSTRACT Salmonella enterica serotype Reading has recently been identified as a significant foodborne pathogen from contaminated poultry products. There is a critical need for close monitoring of this newly emerged pathogen. This study developed bioluminescent strains of S. Reading for real-time pathogen tracking using bioluminescence imaging. Two strains of S. Reading were used: an outbreak strain and a non-outbreak strain. The chloramphenicol acetyltransferase gene was cloned into the plasmid pBS-slpGFPluxABCDE, which carries luxABCDE operon and an ampicillin resistance gene. The newly constructed plasmid was then transformed into the outbreak and non-outbreak strains of S. Reading by electroporation. The resulting colonies were confirmed by visualizing bioluminescence using an in vivo imaging system and by testing their resistance to chloramphenicol. These strains demonstrated a high bioluminescence level (108–109 Photons/s/cm2/sr) and were tested for growth and plasmid stability by daily subculturing in Luria–Bertani medium with and without antibiotics. The plasmid remained stable for 8 days under non-selective conditions, and growth rates were comparable to non-bioluminescent parent strains in antibiotic-free conditions. However, growth was notably different in the presence of chloramphenicol, indicating successful plasmid retention and function. This study successfully created stable bioluminescent S. Reading strains, marking a significant step forward in monitoring and potentially reducing the spread of this emergent foodborne pathogen in the poultry industry.IMPORTANCESalmonella enterica serotype Reading has recently become a significant foodborne pathogen linked to poultry products. To enhance pathogen monitoring, this study developed bioluminescent strains of S. Reading by inserting the chloramphenicol acetyltransferase gene into a plasmid containing a bioluminescence gene cluster. These modified strains were transformed into outbreak and non-outbreak bacterial strains via electroporation. The bioluminescent strains demonstrated stable plasmid retention and high bioluminescence levels. They also showed growth comparable to their parent strains, even in the absence of antibiotics. These bioluminescent strains could potentially facilitate real-time monitoring and control of S. Reading in poultry industries.https://journals.asm.org/doi/10.1128/spectrum.01263-24Salmonella Readingfoodborne pathogenoutbreakstransformationbioluminescence imaging |
| spellingShingle | Abubakar Shitu Isah Reshma Ramachandran Anuraj Theradiyil Sukumaran Aaron S. Kiess Claudia D. Castañeda Timothy Boltz Kenneth S. Macklin Hossam Abdelhamed Li Zhang Construction and characterization of bioluminescent Salmonella Reading outbreak and non-outbreak strains Microbiology Spectrum Salmonella Reading foodborne pathogen outbreaks transformation bioluminescence imaging |
| title | Construction and characterization of bioluminescent Salmonella Reading outbreak and non-outbreak strains |
| title_full | Construction and characterization of bioluminescent Salmonella Reading outbreak and non-outbreak strains |
| title_fullStr | Construction and characterization of bioluminescent Salmonella Reading outbreak and non-outbreak strains |
| title_full_unstemmed | Construction and characterization of bioluminescent Salmonella Reading outbreak and non-outbreak strains |
| title_short | Construction and characterization of bioluminescent Salmonella Reading outbreak and non-outbreak strains |
| title_sort | construction and characterization of bioluminescent salmonella reading outbreak and non outbreak strains |
| topic | Salmonella Reading foodborne pathogen outbreaks transformation bioluminescence imaging |
| url | https://journals.asm.org/doi/10.1128/spectrum.01263-24 |
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