As an arduous clinical challenge, oral infections in humans remain widespread despite continuous efforts from dental health professionals [1]. Oral biofilms composed of multispecies microbial communities are considered to play a critical role in triggering infectious dental diseases, such as caries, pulpal, and periodontal inflammation [2,3]. Aside from the tooth surface, the surface of dental restorations has also been regarded as an important platform for bacterial attachment and biofilm growth [4]. Dental restorations are reported to be found in approximately 85% of North American adults (20–64 years old) in recent studies [5,6]. Higher density of biofilm has been reported on various dental restorative materials, including composite resins, when compared to the enamel surface [7]. Although surface roughness influences early bacterial adhesion on composite resins [8], optimizing the surface roughness may only reduce rather than prevent biofilm formation [9]. These threats may increase the risk of oral infections, resulting in significant damage to natural teeth and supporting tissues as well as the need for frequent replacement of the restoration. Therefore, in the case of high-risk caries patients, it is essential to effectively eliminate oral bacteria residing in biofilms on dental restorative materials while ensuring the maintenance of a healthy oral microbiome to mitigate dysbiosis.

Chemical plaque control is a valid and acceptable principle when used as an adjunct to mechanical oral hygiene [10]. Chlorhexidine rinses have been demonstrated to be both safe and effective and should be utilized for the short and medium term when mechanical tooth cleaning is not feasible, challenging, or insufficient [11]. In particular, high-risk patients require antibacterial therapy to reduce the bacterial burden [12]. Antimicrobial mouthrinses have a wide range of applications and have been shown in clinical and in vivo studies to reduce plaque accumulation and microbial pathogenicity, thereby preventing infection [13], [14], [15]. However, biofilms are highly difficult to treat and are recalcitrant to most antibiotics due to their distinct cell gene expression, existing extracellular polymeric substances (EPS/matrix), and heterogeneous organization in microbial communities [16,17]. The overgrowth of resistant or opportunistic pathogens is likely to disrupt natural oral microbial ecology [18]. As a result, more desirable antibiofilm compounds and strategies to improve efficacy in combating oral infection are urgently needed.

Antibiofilm host defense (antimicrobial) peptides have been suggested for treatment of oral multispecies biofilm-induced infections [19]. Pre-existing biofilms formed by both Gram-negative and Gram-positive bacteria can be both killed and dispersed by antibiofilm peptides, to prevent further biofilm formation that specifically target microbial biofilms [20,21]. Based on immune cell activation and autophagy and apoptosis regulation, reduced inflammation and increased microbial killing may result from immunomodulation [22]. In recent years, DJK-5, a d-enantiomeric cationic peptide, has attracted extensive attention for its broad-spectrum antibiofilm capability in the biomedical field [23,24]. The degradation of the stress alarmone guanosine tetraphosphate (ppGpp) which is vital for biofilm development is promoted when DJK-5 interacts with microorganisms leading to suppression of biofilm accumulation, killing of biofilm cells and biofilm dispersal [20,25] (Scheme 1). Our previous studies have demonstrated that DJK-5 can kill single-species (Enterococcus faecalis) and oral multispecies biofilms grown on hydroxyapatite and dentin substrates [24,26]. These observations indicate that DJK-5 may act as a promising antibiofilm alternative to conventional antimicrobial agents in the long-term treatment of oral infection. However, to date, no information is available about the effectiveness of DJK-5 on multispecies biofilms grown on dental restorative materials.

The present study aimed to establish a multispecies biofilm model on different dental restorative materials to assess the dynamics of killing by the peptide DJK-5 and compare it to conventional mouthrinses. The null hypothesis was that there is no difference in the effectiveness of DJK-5 and other mouthrinses against biofilms grown on five restorative materials during a 1-week application period.