Studies have suggested the need for sterilization or disinfection of materials prior to their administration in the oral cavity [8, 12]. However, the use of orthodontic appliances directly from the manufacturer’s packages is still a routine clinical practice. According to previous studies, orthodontic appliances received from the manufacturer’s packages were unsterile [1, 10, 11]. Therefore, the present study evaluated the bacterial load of the orthodontic brackets received from different manufacturers and determined the efficacy of chlorhexidine in destroying the microbial contamination.
Bacterial colonization was confirmed in all the orthodontic brackets received from different manufacturers. The outcome of the current investigation was similar to prior studies using different orthodontic appliances, such as arch wires , orthodontic pliers , brackets , orthodontic buccal tubes , and tooth brushes  received from different manufacturers. These studies indicate that orthodontic appliances used in dentistry are often contaminated with bacteria.
In our study, Staphylococci were the predominant organisms isolated from orthodontic brackets. Contamination with Staphylococci mostly occurs due to skin contact during manufacturing and/or packaging of orthodontic appliances [1, 6]. Similar studies conducted in this regard reported that Staphylococci were the common organisms to contaminate the orthodontic brackets [1, 3, 14]. In our study, B. cereus and B. licheniformis were the other frequently isolated organisms from the orthodontic brackets, followed by Streptococci. Bacillus spp. cause food-borne diseases as well as nosocomial outbreaks in immune-suppressed hospitalized patients .
K. pneumoniae is the respiratory pathogen that was isolated from orthodontic brackets in our study. The infection spreads from one person to the other through contaminated hands of individuals in the hospital. A similar study conducted by Rastogi et al.  isolated Klebsiella spp. from the orthodontic brackets. Further, literature reported a direct association of Klebsiella spp. with autoimmune disorders, such as ankylosing spondylitis, rheumatoid arthritis, and Crohn’s disease [16, 17]. Isolation of Lactobacilli spp. that initiate and progress dental caries/decay was relatively low in our study . All these potential microorganisms are of major health concern; therefore, it is essential to sterilize or disinfect the brackets before fixing in the oral cavity. The other non-pathogenic bacteria isolated from the brackets in group 2 were not detrimental to the patients’ health.
Chlorhexidine used in various medical fields, such as gynecology, urology, and ophthalmology, has a broad antimicrobial activity . Several studies demonstrated that chlorhexidine is effective both as an antiplaque and antimicrobial agent. Depending on different concentrations, it has both bacteriostatic and bactericidal properties [19, 20]. Research has further reported that chlorhexidine does not affect the shear bond strength of orthodontic brackets and clinically exhibits acceptable bond strength . Speer et al. also reported that chlorhexidine did not affect the bond strength of metal brackets; however, it reduced the bond strength of ceramic brackets . In our study, two concentrations (0.01% and 2%) of chlorhexidine were used to disinfect the orthodontic brackets received from different manufacturers. Initially, the most commonly used 0.01% chlorhexidine, commercially available as mouthwash, was used for disinfection. However, complete disinfection was not observed in all the groups. Due to incomplete disinfection, 2% chlorhexidine solution—the next higher concentration used in the medical field—was used for disinfection. The exact mechanism exerted by chlorhexidine in destroying the bacteria is not yet clear . However, it has been postulated that positively charged chlorhexidine molecules bind to the negatively charged lipid molecules of the cell membrane and interfere with the process of osmosis.  The other novel approach that can be used to reduce the bacterial contamination of orthodontic brackets is application of antimicrobial nanoparticles . The different methods include coating of orthodontic brackets with a thin film of nitrogen-doped titania nanoparticles; combination of glass ionomer or resin-modified glass ionomer cements with fluorapatite, fluorohydroxyapatite, or hydroxyapatite nanoparticles; addition of titania, silica, or silver nanoparticles to acrylic orthodontic materials; and incorporation of nanofillers or silica/titania nanoparticles into orthodontic adhesives .
Studies have demonstrated that slightly higher concentrations of chlorhexidine are required to kill gram-negative pathogens than those required to kill the gram-positive pathogens [24, 25]. Due to the presence of a permeable cell wall in the gram-positive bacteria, they are destroyed easily when compared to the gram-negative bacteria . Organisms present in group 2 were gram-positive and non-pathogenic bacteria. Therefore, a lower concentration (0.01%) of chlorohexidine was adequate to destroy all the bacteria. However, both gram-positive and gram-negative bacteria were observed on brackets in other groups, which required a higher concentration (2%) of chlorhexidine for complete decontamination.
Although unique, the current study has some potential limitations. As the study was conducted in in vitro conditions, further in vivo studies are required to support these findings. While orthodontic brackets showed complete decontamination after treatment with 2% chlorhexidine, there is no data related to long-term effectiveness of chlorhexidine to impede the growth of microorganisms.
Overall, the results advocate that the orthodontic brackets received from the manufacturer require suitable disinfection to safeguard the patients’ health. Furthermore, clinicians should be cautious about the use of contaminated appliances prior to administering in the oral cavity as it might affect the systemic health of the patients.