OxyR inactivation reduces the growth rate and oxidative stress defense in Capnocytophaga ochracea

Abstract

Objective: The human oral cavity harbors several bacteria. Among them, Capnocytophaga ochracea, a facultative anaerobe, is responsible for the early phase of dental plaque formation. In this phase, the tooth surface or tissue is exposed to various oxidative stresses. For colonization in the dental plaque phase, a response by hydrogen peroxide (H₂O₂)-sensing transcriptional regulators, such as OxyR, may be necessary. However, to date, no study has elucidated the role of OxyR protein in C. ochracea.

Methods: Insertional mutagenesis was used to create an oxyR mutant, and gene expression was evaluated by reverse transcription-polymerase chain reaction and quantitative real-time reverse transcription-polymerase chain reaction. Bacterial growth curves were generated by turbidity measurement, and the sensitivity of the oxyR mutant to H₂O₂ was assessed using the disc diffusion assay. Finally, a two-compartment system was used to assess biofilm formation.

Results: The oxyR mutant grew slower than the wild-type under anaerobic conditions. The agar diffusion assay revealed that the oxyR mutant had increased sensitivity to H₂O₂. The transcript levels of oxidative stress defense genes, sod, ahpC, and trx, were lower in the oxyR mutant than in the wild-type strain. The turbidity of C. ochracea, simultaneously co-cultured with Streptococcus gordonii, was lower than that observed under conditions of homotypic growth. Moreover, the percentage decrease in growth of the oxyR mutant was significantly higher than that of the wild-type.

Conclusions: These results show that OxyR in C. ochracea regulates adequate in vitro growth and escapes oxidative stress.

Keywords: Capnocytophaga ochracea; Hydrogen peroxide; In vitro growth; OxyR; Polymicrobial community; Streptococcus gordonii.