Aditya Banerjee

Since model dose-response is critical when assessing caries lesion development over time, this study evaluated the influence of fluoride dose and treatment duration on caries progression in a rat caries model. Streptococcus mutans -infected Sprague-Dawley rats were treated with deionized water, 226 ppm F ⁻ , or 2,260 ppm F ⁻ twice daily for 3, 4, or 5 weeks. Caries lesions were assessed using Larson’s modification of the Keyes scoring system and complemented by micro-computed tomography (µCxT). Intraoral fluoride availability, serum and bone fluoride concentrations and microbial counts were also determined. Fluoride reduced caries severity in a dose- and time-dependent manner. While early enamel lesions were detected in all groups, extensive dentine lesions increased over time, in a dose-dependent manner, in the control and 226 ppm F ⁻ groups, and were not observed in the 2,260 ppm F ⁻ group after 5 weeks. Intraoral and bone fluoride availability increased significantly with fluoride concentration and treatment duration, whereas serum fluoride levels reflected fluoride dose instead of treatment duration. µCT-derived enamel volume correlated negatively with both total and extensive caries scores, supporting its utility as an objective measure of lesion severity. In conclusion, extending model length from 3 to 5 weeks increased the severity of caries lesions in a dose-dependent manner. Fluoride intraoral availability and bone fluoride also demonstrated a dose and time-dependent response.

Rapid growth in the nuclear energy sector has led to increased construction of nuclear power plants (NPPs). Although this promotes the generation of alternate sources of “clean” energy that does not harm the environment, potential concerns regarding soil and water pollution with microplastics and multiple rare heavy metals (HMs) used in NPPs usually do not grab the required attention. The group of rare HMs comprises of uranium, cadmium, mercury, cobalt, germanium, and indium, which are known ecological toxins affecting agricultural quality and consumer safety. When expunged as nuclear waste discharges, these rare HMs adsorb to the surface of microplastics and together pollute the adjacent cultivable lands and water sources used for irrigation. Microplastics increase the phyto-availability of the HMs, which mimic micronutrient elements and are actively transported into root cells via calcium, iron, zinc, copper, or other HM transporters. The toxicants are then translocated to aerial biomass and reproductive or storage organs via the symplastic or apoplastic routes. Humans or animals consuming such contaminated crops and vegetables can develop irreversible neurological and physiological disorders, including cancers. Plant growth regulators like abscisic acid, gibberellic acid, and nitric oxide have been found to synchronize the stress-adaptive signaling in crops, although the sensitive genotypes ultimately succumb to oxidative injuries. To abate such ecological and economic loss, remote sensing can be used to avoid contaminated areas or bio(phyto)remediation can be performed to depollute contaminated landscapes and water bodies. Genetically engineered, tolerant crops can also be cultivated directly, with lower yield loss.

Fluoride has long been known to possess antimicrobial properties. For many bacteria, the toxic effects of fluoride are reversible. However, fluoride has also been shown to trigger lysis and cell death in many other diverse bacterial species, including dental pathogens. The underlying molecular mechanisms responsible for fluoride-induced cell lysis have not been established. Using Streptococcus mutans as a model, we show that fluoride elicits an uncontrolled stress response characterized by upregulation of competence pathways and extensive cell wall degradation. While controlled and limited autolysis under stress is an adaptive response, we show that expression of the competence-associated alternative sigma factor ComX is prolonged under fluoride stress, relative to other stressors. Using in vitro and in vivo analyses, we show that fluoride disrupts the typical tight temporal control of ComX by specifically inhibiting assembly and activity of Clp ATPases responsible for its proteolytic degradation. Unchecked, ComX upregulates bacteriocins and autolysins, while simultaneously suppressing immunity peptide expression via a 6S RNA-mediated mechanism. Thus, fluoride subverts cellular mechanisms to turn off competence pathways that are induced under cellular stress, causing irreversible damage to the cell wall and ultimately cell death. Phenylalanine partially restores Clp protease assembly and activity, providing a rationale for the frequent presence of a gene encoding chorismate mutase in fluoride-responsive operons. Together, our findings reveal the molecular mechanism of fluoride-dependent lysis in bacteria, fifty years after this phenomenon was first reported. These pathways could be exploited to potentiate the antimicrobial effects of oral fluoride.

Fluoride has long been known to possess antimicrobial properties, triggering extensive lysis in diverse bacterial species. Surprisingly, given the ubiquity of fluoride in oral healthcare, the underlying mechanisms by which fluoride kills bacteria remain unknown. Using dental pathogen Streptococcus mutans as a model, we show that fluoride elicits an uncontrolled stress response characterized by upregulation of competence pathways and extensive cell wall degradation. While limited autolysis is adaptive, we show that fluoride subverts the typical tight temporal control of the competence-associated alternative sigma factor ComX by specifically inhibiting assembly and activity of Clp ATPases, thus disrupting proteolytic degradation of ComX. Phenylalanine partially restores Clp protease activity, explaining the frequent presence of a gene encoding chorismate mutase in fluoride-responsive operons. Together, these findings reveal the molecular mechanism that underlie fluoride-dependent lysis in bacteria, and identify new pathways that could be exploited to potentiate the antimicrobial effects of oral fluoride.

Microorganisms resist fluoride toxicity using fluoride export proteins from one of several different molecular families. Cariogenic species Streptococcus mutans and Candida albicans extrude intracellular fluoride using a CLC ^F F ^- /H ⁺ antiporter and FEX fluoride channel, respectively, whereas oral commensal eubacteria, such as Streptococcus gordonii, export fluoride using a Fluc fluoride channel. In this work, we examine how genetic knockout of fluoride export impacts pathogen fitness in single-species and three-species dental biofilm models. For biofilms generated using S. mutans with the genetic knockout of the CLC ^F transporter, exposure to low fluoride concentrations decreased S. mutans counts, synergistically reduced the populations of C. albicans , increased the relative proportion of oral commensal S. gordonii , and reduced properties associated with biofilm pathogenicity, including acid production and hydroxyapatite dissolution. Biofilms prepared with C. albicans with genetic knockout of the FEX channel also exhibited reduced fitness in the presence of fluoride but to a lesser degree. Imaging studies indicate that S. mutans is highly sensitive to fluoride, with the knockout strain undergoing complete lysis when exposed to low fluoride for a moderate amount of time. Biochemical purification of the S. mutans CLC ^F transporter and functional reconstitution establishes that the functional protein is a dimer encoded by a single gene. Together, these findings suggest that fluoride export by oral pathogens can be targeted by specific inhibitors to restore biofilm symbiosis in dental biofilms and that S. mutans is especially susceptible to fluoride toxicity. IMPORTANCE: Dental caries is a globally prevalent condition that occurs when pathogenic species, including Streptococcus mutans and Candida albicans , outcompete beneficial species, such as Streptococcus gordonii , in the dental biofilm. Fluoride is routinely used in oral hygiene to prevent dental caries. Fluoride also has antimicrobial properties, although most microbes possess fluoride exporters to resist its toxicity. This work shows that sensitization of cariogenic species S. mutans and C. albicans to fluoride by genetic knockout of fluoride exporters alters the microbial composition and pathogenic properties of dental biofilms. These results suggest that the development of drugs that inhibit fluoride exporters could potentiate the anticaries effect of fluoride in over-the-counter products like toothpaste and mouth rinses. This is a novel strategy to treat dental caries.

Nanomaterials (NMs) are increasingly being used in plant science to promote plant growth, development, and biofortification. Apart from their uses in genetic engineering, NMs are also used as nanofertilizers, nanosensors, and nanobionics within plant systems. Several studies have proved the utility of NMs in improving plant productivity and nutrition even under sub-optimal and environmentally challenging conditions. In this regard, the chapter serves the purpose to briefly introduce the recent advancements in the use of NMs in advancing plant science research.