Structural basis of cyclic oligoadenylate binding to the transcription factor Csa3 outlines cross talk between type III and type I CRISPR systems.

RNA interference by type III CRISPR systems results in the synthesis of cyclic oligoadenylate (cOA) second messengers, which are known to bind and regulate various CARF domain-containing nuclease receptors. The CARF domain-containing Csa3 family of transcriptional factors associated with the DNA-targeting type I CRISPR systems regulate expression of various CRISPR and DNA repair genes in many prokaryotes. In this study, we extend the known receptor repertoire of cOA messengers to include transcriptional factors by demonstrating specific binding of cyclic tetra-adenylate (cA4) to Saccharolobus solfataricus Csa3 (Csa3Sso). Our 2.0-Å resolution X-ray crystal structure of cA4-bound full-length Csa3Sso reveals the binding of its CARF domain to an elongated conformation of cA4. Using cA4 binding affinity analyses of Csa3Sso mutants targeting the observed Csa3Sso•cA4 structural interface, we identified a Csa3-specific cA4 binding motif distinct from a more widely conserved cOA-binding CARF motif. Using a rational surface engineering approach, we increased the cA4 binding affinity of Csa3Sso up to ∼145-fold over the wildtype, which has potential applications for future second messenger-driven CRISPR gene expression and editing systems. Our in-solution Csa3Sso structural analysis identified cA4-induced allosteric and asymmetric conformational rearrangement of its C-terminal winged helix-turn-helix effector domains, which could potentially be incompatible to DNA binding. However, specific in vitro binding of the purified Csa3Sso to its putative promoter (PCas4a) was found to be cA4 independent, suggesting a complex mode of Csa3Sso regulation. Overall, our results support cA4-and Csa3-mediated cross talk between type III and type I CRISPR systems.

Structural basis of KdpD histidine kinase binding to the second messenger c-di-AMP.

The KdpDE two-component system regulates potassium homeostasis and virulence in various bacterial species. The KdpD histidine kinases (HK) of this system contain a universal stress protein (USP) domain which binds to the second messenger cyclic-di-adenosine monophosphate (c-di-AMP) for regulating transcriptional output from this two-component system in Firmicutes such as Staphylococcus aureus. However, the structural basis of c-di-AMP specificity within the KdpD-USP domain is not well understood. Here, we resolved a 2.3 Å crystal structure of the S. aureus KdpD-USP domain (USPSa) complexed with c-di-AMP. Binding affinity analyses of USPSa mutants targeting the observed USPSa:c-di-AMP structural interface enabled the identification of the sequence residues that are required for c-di-AMP specificity. Based on the conservation of these residues in other Firmicutes, we identified the binding motif, (A/G/C)XSXSX2N(Y/F), which allowed us to predict c-di-AMP binding in other KdpD HKs. Furthermore, we found that the USPSa domain contains structural features distinct from the canonical standalone USPs that bind ATP as a preferred ligand. These features include inward-facing conformations of its ß1-α1 and ß4-α4 loops, a short α2 helix, the absence of a triphosphate-binding Walker A motif, and a unique dual phospho-ligand binding mode. It is therefore likely that USPSa-like domains in KdpD HKs represent a novel subfamily of the USPs.

Frequency and types of incidental findings on limited field of view CBCT scans.

The objective of this retrospective study was to identify and categorize incidental findings (IFs) discovered on limited field of view (LFOV) cone beam computed tomographic (CBCT) scans of the maxillofacial region. The sequential LFOV CBCT radiology reports created by 2 board-certified radiologists from January 1, 2017, to December 31, 2017, were retrospectively reviewed. The CBCT scans were acquired for implant site evaluation. Incidental findings of pathologic or anatomical defects were categorized as endodontic, periodontal, sinus-related, nonodontogenic, or other concerns. Of 474 radiology reports that were evaluated, 244 (51.5%) recorded at least 1 detectable IF. Fifty-one of these 244 (10.8% of the total sample) recorded that multiple IFs were detected on a single CBCT scan. On 3 reports, 3 IFs were detected, while the remaining 48 reports showed 2 IFs. More than half of the 295 unique IFs were categorized as endodontic or sinus-related concerns. The majority (70.5%) of IFs were recommended for referral or additional treatment. The results indicate that the use of a CBCT system with an LFOV does not diminish the need to conduct a thorough evaluation of the images.

Investigations of Dimethylglycine, Glycine Betaine, and Ectoine Uptake by a Betaine-Carnitine-Choline Transporter Family Transporter with Diverse Substrate Specificity in Vibrio Species.

Fluctuations in osmolarity are one of the most prevalent stresses to which bacteria must adapt, both hypo- and hyperosmotic conditions. Most bacteria cope with high osmolarity by accumulating compatible solutes (osmolytes) in the cytoplasm to maintain the turgor pressure of the cell. Vibrio parahaemolyticus, a halophile, utilizes at least six compatible solute transporters for the uptake of osmolytes: two ABC family ProU transporters and four betaine-carnitine-choline transporter (BCCT) family transporters. The full range of compatible solutes transported by this species has yet to be determined. Using an osmolyte phenotypic microarray plate for growth analyses, we expanded the known osmolytes used by V. parahaemolyticus to include N,N-dimethylglycine (DMG), among others. Growth pattern analysis of four triple-bccT mutants, possessing only one functional BCCT, indicated that BccT1 (VP1456), BccT2 (VP1723), and BccT3 (VP1905) transported DMG. BccT1 was unusual in that it could take up both compounds with methylated head groups (glycine betaine [GB], choline, and DMG) and cyclic compounds (ectoine and proline). Bioinformatics analysis identified the four coordinating amino acid residues for GB in the BccT1 protein. In silico modeling analysis demonstrated that GB, DMG, and ectoine docked in the same binding pocket in BccT1. Using site-directed mutagenesis, we showed that a strain with all four residues mutated resulted in the loss of uptake of GB, DMG, and ectoine. We showed that three of the four residues were essential for ectoine uptake, whereas only one of the residues was important for GB uptake. Overall, we have demonstrated that DMG is a highly effective compatible solute for Vibrio species and have elucidated the amino acid residues in BccT1 that are important for the coordination of GB, DMG, and ectoine transport.IMPORTANCEVibrio parahaemolyticus possesses at least six osmolyte transporters, which allow the bacterium to adapt to high-salinity conditions. In this study, we identified several additional osmolytes that were utilized by V. parahaemolyticus We demonstrated that the compound DMG, which is present in the marine environment, was a highly effective osmolyte for Vibrio species. We determined that DMG is transported via BCCT family carriers, which have not been shown previously to take up this compound. BccT1 was a carrier for GB, DMG, and ectoine, and we identified the amino acid residues essential for the coordination of these compounds. The data suggest that for BccT1, GB is more easily accommodated than ectoine in the transporter binding pocket.

Effect of Intracanal Medicaments (Modified Triple Antibiotic Paste, Calcium Hydroxide, and Aloe Vera) on Microhardness of Root Dentine: An In Vitro Study.

AIM: To compare the effect of three different intracanal medicaments, namely, modified triple antibiotic paste (MTAP), calcium hydroxide (Ca(OH)2), and aloe vera, on the root dentine microhardness. MATERIALS AND METHODS: A total of 50 extracted mandibular bicuspids were prepared using ProTaper Next rotary files. The roots of the bicuspids were alienated to three groups (n = 10 each) and one control group (untreated; n = 20). In three groups, the root canals were filled with MTAP, Ca(OH)2, and aloe vera medicaments. After 21 days, medicaments were removed by Endo activator. Mean Knoop hardness numbers were calculated after treatment and compared with the untreated control group. Data were evaluated using the Student's t test (paired), ANOVA (one-way) followed, and the post hoc test. RESULTS: All treated groups except the aloe vera group had shown significant reduction (p < 0.05) in microhardness of the root dentin as compared with the untreated control group. The aloe vera group showed least reduction of microhardness and was statistically insignificant (p > 0.05). CONCLUSION: Aloe vera shows promising results in terms of fewer effects on microhardness of the root dentin compared to MTAP and Ca(OH)2. CLINICAL SIGNIFICANCE: Elimination of most of the bacterial infection from the root canal and very minimum to no effect on the microhardness of the dentin in the root part are the basics of success in any endodontic treatment. Further in vivo studies are required to compare the efficacy of these intracanal medicaments.

Regenerative Evaluation of Immature Roots using PRF and Artificial Scaffolds in Necrotic Permanent Teeth: A Clinical Study.

AIM: The aim of this study is to evaluate and compare the regenerative potential of natural scaffold [platelet-rich fibrin (PRF)] and artificial scaffolds (commercially available collagen, placentrex, and chitosan) in necrotic immature permanent teeth. MATERIALS AND METHODS: Necrotic immature permanent maxillary incisors with or without radiographic evidence of periapical lesion were included. Access opening was done under rubber dam isolation. Canal disinfection was done using minimal instrumentation, copious irrigation, and double antibiotic paste as interappointment medicament for 4 weeks. After 4 weeks, asymptomatic teeth were divided into four groups on the basis of scaffolds used for the revascularization procedure: group I (PRF); group II (collagen); group III (placentrex); group IV (chitosan). The clinical and radiographic evaluations of teeth were done at 3, 6, and 12 months after the procedure and compared with baseline records. RESULTS: Clinically, patients were completely asymptomatic throughout the study period. Radiographically, all cases showed an improvement in terms of periapical healing, apical closure, root lengthening, and dentinal wall thickening. PRF and collagen gave better results than placentrex and chitosan in terms of periapical healing, apical closure, and dentinal wall thickening. CONCLUSION: Revascularization procedure is more effective and conservative over apexification in the management of necrotic immature permanent teeth. This study has shown that PRF and collagen are better scaffolds than placentrex and chitosan for inducing apexogenesis in immature necrotic permanent teeth.

Rgg protein structure-function and inhibition by cyclic peptide compounds.

Peptide pheromone cell-cell signaling (quorum sensing) regulates the expression of diverse developmental phenotypes (including virulence) in Firmicutes, which includes common human pathogens, e.g., Streptococcus pyogenes and Streptococcus pneumoniae. Cytoplasmic transcription factors known as "Rgg proteins" are peptide pheromone receptors ubiquitous in Firmicutes. Here we present X-ray crystal structures of a Streptococcus Rgg protein alone and in complex with a tight-binding signaling antagonist, the cyclic undecapeptide cyclosporin A. To our knowledge, these represent the first Rgg protein X-ray crystal structures. Based on the results of extensive structure-function analysis, we reveal the peptide pheromone-binding site and the mechanism by which cyclosporin A inhibits activation of the peptide pheromone receptor. Guided by the Rgg-cyclosporin A complex structure, we predicted that the nonimmunosuppressive cyclosporin A analog valspodar would inhibit Rgg activation. Indeed, we found that, like cyclosporin A, valspodar inhibits peptide pheromone activation of conserved Rgg proteins in medically relevant Streptococcus species. Finally, the crystal structures presented here revealed that the Rgg protein DNA-binding domains are covalently linked across their dimerization interface by a disulfide bond formed by a highly conserved cysteine. The DNA-binding domain dimerization interface observed in our structures is essentially identical to the interfaces previously described for other members of the XRE DNA-binding domain family, but the presence of an intermolecular disulfide bond buried in this interface appears to be unique. We hypothesize that this disulfide bond may, under the right conditions, affect Rgg monomer-dimer equilibrium, stabilize Rgg conformation, or serve as a redox-sensitive switch.

Conformational change-induced repeat domain expansion regulates Rap phosphatase quorum-sensing signal receptors.

The large family of Gram-positive quorum-sensing receptors known as the RNPP proteins consists of receptors homologous to the Rap, NprR, PlcR, and PrgX proteins that are regulated by imported oligopeptide autoinducers. Rap proteins are phosphatases and transcriptional anti-activators, and NprR, PlcR, and PrgX proteins are DNA binding transcription factors. Despite their obvious importance, the mechanistic basis of oligopeptide receptor regulation is largely unknown. Here, we report the X-ray crystal structure of the Bacillus subtilis quorum-sensing receptor RapJ in complex with the centrally important oligopeptide autoinducer competence and sporulation factor (CSF, also termed PhrC), a member of the Phr family of quorum-sensing signals. Furthermore, we present the crystal structure of RapI. Comparison of the RapJ-PhrC, RapI, RapH-Spo0F, and RapF-ComA(C) crystal structures reveals the mechanistic basis of Phr activity. More specifically, when complexed with target proteins, Rap proteins consist of a C-terminal tetratricopeptide repeat (TPR) domain connected by a flexible helix-containing linker to an N-terminal 3-helix bundle. In the absence of a target protein or regulatory peptide, the Rap protein 3-helix bundle adopts different conformations. However, in the peptide-bound conformation, the Rap protein N-terminal 3-helix bundle and linker undergo a radical conformational change, form TPR-like folds, and merge with the existing C-terminal TPR domain. To our knowledge, this is the first example of conformational change-induced repeat domain expansion. Furthermore, upon Phr binding, the entire Rap protein is compressed along the TPR superhelical axis, generating new intramolecular contacts that lock the Rap protein in an inactive state. The fact that Rap proteins are conformationally flexible is surprising considering that it is accepted dogma that TPR proteins do not undergo large conformational changes. Repeat proteins are widely used as scaffolds for the development of designed affinity reagents, and we propose that Rap proteins could be used as scaffolds for engineering novel ligand-switchable affinity reagents.

Antituberculosis thiophenes define a requirement for Pks13 in mycolic acid biosynthesis.

We report a new class of thiophene (TP) compounds that kill Mycobacterium tuberculosis by the previously uncharacterized mechanism of Pks13 inhibition. An F79S mutation near the catalytic Ser55 site in Pks13 conferred TP resistance in M. tuberculosis. Overexpression of wild-type Pks13 resulted in TP resistance, and overexpression of the Pks13(F79S) mutant conferred high resistance. In vitro, TP inhibited fatty acyl-AMP loading onto Pks13. TP inhibited mycolic acid biosynthesis in wild-type M. tuberculosis, but it did so to a much lesser extent in TP-resistant M. tuberculosis. TP treatment was bactericidal and equivalent to treatment with the first-line drug isoniazid, but it was less likely to permit emergent resistance. Combined isoniazid and TP treatment resulted in sterilizing activity. Computational docking identified a possible TP-binding groove within the Pks13 acyl carrier protein domain. This study confirms that M. tuberculosis Pks13 is required for mycolic acid biosynthesis, validates it as a druggable target and demonstrates the therapeutic potential of simultaneously inhibiting multiple targets in the same biosynthetic pathway.

Comparison of state dental radiography safety regulations.

The aim of this study was to compare and provide an overview of state policies on occupational exposure, dosimetry, collimation, patient protection, and the use of portable handheld X-ray machines in dentistry. State government webpages containing radiation protection rules and regulations were scanned. The contents were compared against current federal regulations established by the Nuclear Regulatory Commission (NRC) and the US Food and Drug Administration (FDA). They were further evaluated in light of current recommendations from the National Council on Radiation Protection & Measurements (NCRP) and the American Dental Association (ADA). Most states' regulations mirror the exposure limits set forth by the NRC and FDA. Nonregulatory recommendations regarding use of dental radiography are periodically put forth by the NCRP and the ADA. State and federal agencies often follow recommendations from these scientific organizations when creating regulations. Clinicians must be aware of their state's radiation protection rules, as variations among states exist. In addition, recommendations published by organizations such as the NCRP and the ADA, while not legally binding, contribute significantly to the reduction of radiation risks for operators and patients alike.

Structural basis of response regulator dephosphorylation by Rap phosphatases.

Bacterial Rap family proteins have been most extensively studied in Bacillus subtilis, where they regulate activities including sporulation, genetic competence, antibiotic expression, and the movement of the ICEBs1 transposon. One subset of Rap proteins consists of phosphatases that control B. subtilis and B. anthracis sporulation by dephosphorylating the response regulator Spo0F. The mechanistic basis of Rap phosphatase activity was unknown. Here we present the RapH-Spo0F X-ray crystal structure, which shows that Rap proteins consist of a 3-helix bundle and a tetratricopeptide repeat domain. Extensive biochemical and genetic functional studies reveal the importance of the observed RapH-Spo0F interactions, including the catalytic role of a glutamine in the RapH 3-helix bundle that inserts into the Spo0F active site. We show that in addition to dephosphorylating Spo0F, RapH can antagonize sporulation by sterically blocking phosphoryl transfer to and from Spo0F. Our structure-function analysis of the RapH-Spo0F interaction identified Rap protein residues critical for Spo0F phosphatase activity. This information enabled us to assign Spo0F phosphatase activity to a Rap protein based on sequence alone, which was not previously possible. Finally, as the ultimate test of our newfound understanding of the structural requirements for Rap phosphatase function, a non-phosphatase Rap protein that inhibits the binding of the response regulator ComA to DNA was rationally engineered to dephosphorylate Spo0F. In addition to revealing the mechanistic basis of response regulator dephosphorylation by Rap proteins, our studies support the previously proposed T-loop-Y allostery model of receiver domain regulation that restricts the aromatic "switch" residue to an internal position when the ß4-α4 loop adopts an active-site proximal conformation.

A plasmid-encoded phosphatase regulates Bacillus subtilis biofilm architecture, sporulation, and genetic competence.

Bacillus subtilis biofilm formation is tightly regulated by elaborate signaling pathways. In contrast to domesticated lab strains of B. subtilis which form smooth, essentially featureless colonies, undomesticated strains such as NCIB 3610 form architecturally complex biofilms. NCIB 3610 also contains an 80-kb plasmid absent from laboratory strains, and mutations in a plasmid-encoded homolog of a Rap protein, RapP, caused a hyperrugose biofilm phenotype. Here we explored the role of rapP phrP in biofilm formation. We found that RapP is a phosphatase that dephosphorylates the intermediate response regulator Spo0F. RapP appears to employ a catalytic glutamate to dephosphorylate the Spo0F aspartyl phosphate, and the implications of the RapP catalytic glutamate are discussed. In addition to regulating B. subtilis biofilm formation, we found that RapP regulates sporulation and genetic competence as a result of its ability to dephosphorylate Spo0F. Interestingly, while rap phr gene cassettes routinely form regulatory pairs; i.e., the mature phr gene product inhibits the activity of the rap gene product, the phrP gene product did not inhibit RapP activity in our assays. RapP activity was, however, inhibited by PhrH in vivo but not in vitro. Additional genetic analysis suggests that RapP is directly inhibited by peptide binding. We speculate that PhrH could be subject to posttranslational modification in vivo and directly inhibit RapP activity or, more likely, PhrH upregulates the expression of a peptide that, in turn, directly binds to RapP and inhibits its Spo0F phosphatase activity.

Unilateral Buccal Bifurcation Cyst: A Rare Cystic Lesion in Children and Adolescents.

A buccal bifurcation cyst (BBC) is a rarely occurring, distinct lesion that is limited exclusively to the buccal bifurcation area of mandibular first and second molars in children and adolescents. A definitive diagnosis is formulated based on specific clinical and radiographic features. Management of such cysts depends on the presence of symptoms and the size of the lesion. This case report details the common features of a BBC in a 13-year-old patient and outlines the surgical approach to managing the cystic entity. The importance of a comprehensive clinical examination and appropriate supplemental investigations is emphasized to facilitate accurate diagnosis.

Diagnosing Oral Granulomatous Lesions: A Case Report.

The diagnosis of oral granulomatous lesions raises many challenges for the clinician. This article, which includes a case report, describes a process to formulate differential diagnoses by identifying distinguishing characteristics of an entity and applying that information to attain understanding of the ongoing pathophysiological process. Relevant clinical, radiographic, and histologic findings of common disease entities that can mimic clinical and radiographic presentation of this case are discussed to aid dental clinicians in identifying and diagnosing similar lesions in their practice.

Identification of small molecules that antagonize diguanylate cyclase enzymes to inhibit biofilm formation.

Bacterial biofilm formation is responsible for numerous chronic infections, causing a severe health burden. Many of these infections cannot be resolved, as bacteria in biofilms are resistant to the host's immune defenses and antibiotic therapy. New strategies to treat biofilm-based infections are critically needed. Cyclic di-GMP (c-di-GMP) is a widely conserved second-messenger signal essential for biofilm formation. As this signaling system is found only in bacteria, it is an attractive target for the development of new antibiofilm interventions. Here, we describe the results of a high-throughput screen to identify small-molecule inhibitors of diguanylate cyclase (DGC) enzymes that synthesize c-di-GMP. We report seven small molecules that antagonize these enzymes and inhibit biofilm formation by Vibrio cholerae. Moreover, two of these compounds significantly reduce the total concentration of c-di-GMP in V. cholerae, one of which also inhibits biofilm formation by Pseudomonas aeruginosa in a continuous-flow system. These molecules represent the first compounds described that are able to inhibit DGC activity to prevent biofilm formation.

Acute cervicofacial necrotizing fasciitis: three clinical cases and a review of the current literature.

Cervicofacial necrotizing fasciitis (NF) is a rare condition that can quickly become life-threatening if appropriate treatment is delayed. This condition is observed as a rapidly progressive infection that causes extensive necrosis of the superficial fascia. This report presents a case of cervicofacial NF with microbiological isolation of methicillin-resistant Staphylococcus aureus in a patient with a history of uncontrolled diabetes mellitus following a minor scalp trauma. The article also presents two cases of NF secondary to odontogenic infection. The radiographic finding of the presence of gas locules in the facial planes on the CT scan helped to confirm the diagnosis. Patients were treated with broad-spectrum antibiotic therapy, extensive surgical drainage, debridement, and supportive care. Awareness in the dental community of the signs of NF will facilitate optimal patient management.

CircFISH: A Novel Method for the Simultaneous Imaging of Linear and Circular RNAs.

Circular RNAs (circRNAs) are regulatory RNAs which have recently been shown to have clinical significance in several diseases, including, but not limited to, various cancers, neurological diseases and cardiovascular diseases. The function of such regulatory RNAs is largely dependent on their subcellular localization. Several circRNAs have been shown to conduct antagonistic roles compared to the products of the linear isoforms, and thus need to be characterized distinctly from the linear RNAs. However, conventional fluorescent in situ hybridization (FISH) techniques cannot be employed directly to distinguish the signals from linear and circular isoforms because most circRNAs share the same sequence with the linear RNAs. In order to address this unmet need, we adapted the well-established method of single-molecule FISH by designing two sets of probes to differentiate the linear and circular RNA isoforms by virtue of signal colocalization. We call this method 'circular fluorescent in situ hybridization' (circFISH). Linear and circular RNAs were successfully visualized and quantified at a single-molecule resolution in fixed cells. RNase R treatment during the circFISH reduced the levels of linear RNAs while the circRNA levels remain unaltered. Furthermore, cells with shRNAs specific to circRNA showed the loss of circRNA levels, whereas the linear RNA levels were unaffected. The optimization of the in-situ RNase R treatment allowed the multiplexing of circFISH to combine it with organelle staining. CircFISH was found to be compatible with multiple sample types, including cultured cells and fresh-frozen and formalin-fixed tissue sections. Thus, we present circFISH as a versatile method for the simultaneous visualization and quantification of the distribution and localization of linear and circular RNA in fixed cells and tissue samples.

An atypical Phr peptide regulates the developmental switch protein RapH.

Under conditions of nutrient limitation and high population density, the bacterium Bacillus subtilis can initiate a variety of developmental pathways. The signaling systems regulating B. subtilis differentiation are tightly controlled by switch proteins called Raps, named after the founding members of the family, which were shown to be response regulator aspartate phosphatases. A phr gene encoding a secreted pentapeptide that regulates the activity of its associated Rap protein was previously identified downstream of 8 of the chromosomally encoded rap genes. We identify and validate here the sequence of an atypical Phr peptide, PhrH, by in vivo and in vitro analyses. Using a luciferase reporter bioassay combined with in vitro experiments, we found that PhrH is a hexapeptide (TDRNTT), in contrast to the other characterized Phr pentapeptides. We also determined that phrH expression is driven by a promoter lying within rapH. Unlike the previously identified dedicated σ(H)-driven phr promoters, it appears that phrH expression most likely requires σ(A). Furthermore, we show that PhrH can antagonize both of the known activities of RapH: the dephosphorylation of Spo0F and the sequestration of ComA, thus promoting the development of spores and the competent state. Finally, we propose that PhrH is the prototype of a newly identified class of Phr signaling molecules consisting of six amino acids. This class likely includes PhrI, which regulates RapI and the expression, excision, and transfer of the mobile genetic element ICEBs1.

Identification of a novel benzimidazole that inhibits bacterial biofilm formation in a broad-spectrum manner.

Bacterial biofilm formation causes significant industrial economic loss and high morbidity and mortality in medical settings. Biofilms are defined as multicellular communities of bacteria encased in a matrix of protective extracellular polymers. Because biofilms have a high tolerance for treatment with antimicrobials, protect bacteria from immune defense, and resist clearance with standard sanitation protocols, it is critical to develop new approaches to prevent biofilm formation. Here, a novel benzimidazole molecule, named antibiofilm compound 1 (ABC-1), identified in a small-molecule screen, was found to prevent bacterial biofilm formation in multiple Gram-negative and Gram-positive bacterial pathogens, including Pseudomonas aeruginosa and Staphylococcus aureus, on a variety of different surface types. Importantly, ABC-1 itself does not inhibit the growth of bacteria, and it is effective at nanomolar concentrations. Also, coating a polystyrene surface with ABC-1 reduces biofilm formation. These data suggest ABC-1 is a new chemical scaffold for the development of antibiofilm compounds.