Formaldehyde effects on kanamycin resistance gene of inactivated recombinant Escherichia coli vaccines



Earlier studies have demonstrated the use of inactivated recombinant E. coli (bacterins), to protect against Clostridium spp. in vaccinated animals. These bacterins have a simpler, safer, and faster production process. However, these bacterins carry expression plasmids, containing antibiotic resistance gene, which could be assimilate accidentally by environmental microorganisms. Considering this, we aimed to impair this plasmids using formaldehyde at different concentrations.


This compound inactivated the highest density of cells in 24 h. KanR cassette amplification was found to be impaired with 0.8% for 24 h or 0.4% for 72 h. Upon electroporation, E. coli DH5α ultracompetent cells were unable to acquire the plasmids extracted from the bacterins after inactivation procedure. Formaldehyde-treated bacterins were incubated with other viable strains of E. coli, leading to no detectable gene transfer.


We found that this compound is effective as an inactivation agent. Here we demonstrate the biosafety involving antibiotic resistance gene of recombinant E. coli vaccines allowing to industrial production and animal application.

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We would like to thank all the researchers and staff of the Applied Immunology Laboratory (UFPel, Pelotas, RS, Brazil), and to the funding agencies Coordenação de Aperfeiçoamento de Pessoal de Nível Superior – Brasil (CAPES)–Finance Code 001, Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq), and Fundação de Amparo à Pesquisa do Estado do Rio Grande do Sul (FAPERGS).

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RD, EM, LS and RR performed the experiments. RD, MF, GM, AM and FC supervised the study and reviewed of manuscript. RD,MF, CM-J, GM, AM and FC designed the experiments, analysed and interpreted data, and wrote the manuscript.

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Correspondence to Rafael A. Donassolo.

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Donassolo, R.A., Ferreira, M.R.A., Moreira Jr, C. et al. Formaldehyde effects on kanamycin resistance gene of inactivated recombinant Escherichia coli vaccines. Biotechnol Lett (2020).

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  • Antibiotic resistance gene
  • Chemical inactivation
  • Recombinant vaccines
  • Selection marker