Digital Single-operator Cholangioscopy (DSOC) Improves Interobserver Agreement (IOA) and Accuracy for Evaluation of Indeterminate Biliary Strictures: The Monaco Classification.

BACKGROUND: Visual characteristics seen during digital single-operator cholangioscopy (DSOC) have not been validated. The aim of this 2-phase study was to define terminology by consensus for the visual diagnosis of biliary lesions to develop a model for optimization of the diagnostic performance of DSOC. MATERIALS AND METHODS: In phase 1 (criteria identification), video-cholangioscopy clips were reviewed by 12 expert biliary endoscopists, who were blinded to the final diagnosis. Visual criteria were consolidated into the following categories: (1) stricture, (2) lesion, (3) mucosal features, (4) papillary projections, (5) ulceration, (6) abnormal vessels, (7) scarring, (8) pronounced pit pattern.During the second phase (validation), 14 expert endoscopists reviewed DSOC (SpyGlass DS, Boston Scientific) clips using the 8 criteria to assess interobserver agreement (IOA) rate. RESULTS: In phase 1, consensus for visual findings were categorized into 8 criteria titled the "Monaco Classification." The frequency of criteria were: (1) presence of stricture-75%, (2) presence of lesion type-55%, (3) mucosal features-55%, (4) papillary projections-45%, (5) ulceration-42.5%, (6) abnormal vessels-10%, (7) scarring-40%, and (8) pronounced pit pattern-10%. The accuracy on final diagnosis based on visual impression alone was 70%.In phase 2, the IOA rate using Monaco Classification criteria ranged from slight to fair. The presumptive diagnosis IOA was fair (κ=0.31, SE=0.02), and overall diagnostic accuracy was 70%. CONCLUSIONS: The Monaco classification identifies 8 visual criteria for biliary lesions on single-operator digital cholangioscopy. Using the criteria, the IOA and diagnostic accuracy rate of DSOC is improved compared with prior studies.

Toward a Mobile Platform for Real-world Digital Measurement of Depression: User-Centered Design, Data Quality, and Behavioral and Clinical Modeling.

BACKGROUND: Although effective mental health treatments exist, the ability to match individuals to optimal treatments is poor, and timely assessment of response is difficult. One reason for these challenges is the lack of objective measurement of psychiatric symptoms. Sensors and active tasks recorded by smartphones provide a low-burden, low-cost, and scalable way to capture real-world data from patients that could augment clinical decision-making and move the field of mental health closer to measurement-based care. OBJECTIVE: This study tests the feasibility of a fully remote study on individuals with self-reported depression using an Android-based smartphone app to collect subjective and objective measures associated with depression severity. The goals of this pilot study are to develop an engaging user interface for high task adherence through user-centered design; test the quality of collected data from passive sensors; start building clinically relevant behavioral measures (features) from passive sensors and active inputs; and preliminarily explore connections between these features and depression severity. METHODS: A total of 600 participants were asked to download the study app to join this fully remote, observational 12-week study. The app passively collected 20 sensor data streams (eg, ambient audio level, location, and inertial measurement units), and participants were asked to complete daily survey tasks, weekly voice diaries, and the clinically validated Patient Health Questionnaire (PHQ-9) self-survey. Pairwise correlations between derived behavioral features (eg, weekly minutes spent at home) and PHQ-9 were computed. Using these behavioral features, we also constructed an elastic net penalized multivariate logistic regression model predicting depressed versus nondepressed PHQ-9 scores (ie, dichotomized PHQ-9). RESULTS: A total of 415 individuals logged into the app. Over the course of the 12-week study, these participants completed 83.35% (4151/4980) of the PHQ-9s. Applying data sufficiency rules for minimally necessary daily and weekly data resulted in 3779 participant-weeks of data across 384 participants. Using a subset of 34 behavioral features, we found that 11 features showed a significant (P<.001 Benjamini-Hochberg adjusted) Spearman correlation with weekly PHQ-9, including voice diary-derived word sentiment and ambient audio levels. Restricting the data to those cases in which all 34 behavioral features were present, we had available 1013 participant-weeks from 186 participants. The logistic regression model predicting depression status resulted in a 10-fold cross-validated mean area under the curve of 0.656 (SD 0.079). CONCLUSIONS: This study finds a strong proof of concept for the use of a smartphone-based assessment of depression outcomes. Behavioral features derived from passive sensors and active tasks show promising correlations with a validated clinical measure of depression (PHQ-9). Future work is needed to increase scale that may permit the construction of more complex (eg, nonlinear) predictive models and better handle data missingness.

Secondary structure of the mRNA encoding listeriolysin O is essential to establish the replicative niche of L. monocytogenes.

Intracellular pathogens are responsible for an enormous amount of worldwide morbidity and mortality, and each has evolved specialized strategies to establish and maintain their replicative niche. Listeria monocytogenes is a facultative intracellular pathogen that secretes a pore-forming cytolysin called listeriolysin O (LLO), which disrupts the phagosomal membrane and, thereby, allows the bacteria access to their replicative niche in the cytosol. Nonsynonymous and synonymous mutations in a PEST-like domain near the LLO N terminus cause enhanced LLO translation during intracellular growth, leading to host cell death and loss of virulence. Here, we explore the mechanism of translational control and show that there is extensive codon restriction within the PEST-encoding region of the LLO messenger RNA (mRNA) (hly). This region has considerable complementarity with the 5' UTR and is predicted to form an extensive secondary structure that overlaps the ribosome binding site. Analysis of both 5' UTR and synonymous mutations in the PEST-like domain that are predicted to disrupt the secondary structure resulted in up to a 10,000-fold drop in virulence during mouse infection, while compensatory double mutants restored virulence to WT levels. We showed by dynamic protein radiolabeling that LLO synthesis was growth phase-dependent. These data provide a mechanism to explain how the bacteria regulate translation of LLO to promote translation during starvation in a phagosome while repressing it during growth in the cytosol. These studies also provide a molecular explanation for codon bias at the 5' end of this essential determinant of pathogenesis.

(p)ppGpp and c-di-AMP Homeostasis Is Controlled by CbpB in Listeria monocytogenes.

The facultative intracellular pathogen Listeria monocytogenes, like many related Firmicutes, uses the nucleotide second messenger cyclic di-AMP (c-di-AMP) to adapt to changes in nutrient availability, osmotic stress, and the presence of cell wall-acting antibiotics. In rich medium, c-di-AMP is essential; however, mutations in cbpB, the gene encoding c-di-AMP binding protein B, suppress essentiality. In this study, we identified that the reason for cbpB-dependent essentiality is through induction of the stringent response by RelA. RelA is a bifunctional RelA/SpoT homolog (RSH) that modulates levels of (p)ppGpp, a secondary messenger that orchestrates the stringent response through multiple allosteric interactions. We performed a forward genetic suppressor screen on bacteria lacking c-di-AMP to identify genomic mutations that rescued growth while cbpB was constitutively expressed and identified mutations in the synthetase domain of RelA. The synthetase domain of RelA was also identified as an interacting partner of CbpB in a yeast-2-hybrid screen. Biochemical analyses confirmed that free CbpB activates RelA while c-di-AMP inhibits its activation. We solved the crystal structure of CbpB bound and unbound to c-di-AMP and provide insight into the region important for c-di-AMP binding and RelA activation. The results of this study show that CbpB completes a homeostatic regulatory circuit between c-di-AMP and (p)ppGpp in Listeria monocytogenesIMPORTANCE Bacteria must efficiently maintain homeostasis of essential molecules to survive in the environment. We found that the levels of c-di-AMP and (p)ppGpp, two nucleotide second messengers that are highly conserved throughout the microbial world, coexist in a homeostatic loop in the facultative intracellular pathogen Listeria monocytogenes Here, we found that cyclic di-AMP binding protein B (CbpB) acts as a c-di-AMP sensor that promotes the synthesis of (p)ppGpp by binding to RelA when c-di-AMP levels are low. Addition of c-di-AMP prevented RelA activation by binding and sequestering CbpB. Previous studies showed that (p)ppGpp binds and inhibits c-di-AMP phosphodiesterases, resulting in an increase in c-di-AMP. This pathway is controlled via direct enzymatic regulation and indicates an additional mechanism of ribosome-independent stringent activation.

Listeriolysin O: A phagosome-specific cytolysin revisited.

Listeriolysin O (LLO) is an essential determinant of Listeria monocytogenes pathogenesis that mediates the escape of L. monocytogenes from host cell vacuoles, thereby allowing replication in the cytosol without causing appreciable cell death. As a member of the cholesterol-dependent cytolysin (CDC) family of pore-forming toxins, LLO is unique in that it is secreted by a facultative intracellular pathogen, whereas all other CDCs are produced by pathogens that are largely extracellular. Replacement of LLO with other CDCs results in strains that are extremely cytotoxic and 10,000-fold less virulent in mice. LLO has structural and regulatory features that allow it to function intracellularly without causing cell death, most of which map to a unique N-terminal region of LLO referred to as the proline, glutamic acid, serine, threonine (PEST)-like sequence. Yet, while LLO has unique properties required for its intracellular site of action, extracellular LLO, like other CDCs, affects cells in a myriad of ways. Because all CDCs form pores in cholesterol-containing membranes that lead to rapid Ca2+ influx and K+ efflux, they consequently trigger a wide range of host cell responses, including mitogen-activated protein kinase activation, histone modification, and caspase-1 activation. There is no debate that extracellular LLO, like all other CDCs, can stimulate multiple cellular activities, but the primary question we wish to address in this perspective is whether these activities contribute to L. monocytogenes pathogenesis.

Biomechanical Assessment of the Dorsal Spanning Bridge Plate in Distal Radius Fracture Fixation: Implications for Immediate Weight-Bearing.

BACKGROUND: The goal of this study was to compare the biomechanical stability of a 2.4-mm dorsal spanning bridge plate with a volar locking plate (VLP) in a distal radius fracture model, during simulated crutch weight-bearing. METHODS: Five paired cadaveric forearms were tested. A 1-cm dorsal wedge osteotomy was created to simulate an unstable distal radius fracture with dorsal comminution. Fractures were fixed with a VLP or a dorsal bridge plate (DBP). Specimens were mounted to a crutch handle, and optical motion-tracking sensors were attached to the proximal and distal segments. Specimens were loaded in compression at 1 mm/s on a servohydraulic test frame until failure, defined as 2 mm of gap site displacement. RESULTS: The VLP construct was significantly more stable to axial load in a crutch weight-bearing model compared with the DBP plate (VLP: 493 N vs DBP: 332 N). Stiffness was higher in the VLP constructs, but this was not statistically significant (VLP: 51.4 N/mm vs DBP: 32.4 N/mm). With the crutch weight-bearing model, DBP failed consistently with wrist flexion and plate bending, whereas VLP failed with axial compression at the fracture site and dorsal collapse. CONCLUSIONS: Dorsal spanning bridge plating is effective as an internal spanning fixator in treating highly comminuted intra-articular distal radius fracture and prevents axial collapse at the radiocarpal joint. However, bridge plating may not offer advantages in early weight-bearing or transfer in polytrauma patients, with less axial stability in our crutch weight-bearing model compared with volar plating. A stiffer 3.5-mm DBP or use of a DBP construct without the central holes may be considered for distal radius fractures if the goal is early crutch weight-bearing through the injured extremity.

Transforming Psychiatry into Data-Driven Medicine with Digital Measurement Tools.

Psychiatry has been limited by historically rooted practices centered primarily on subjective observation. Fields such as oncology have progressed toward data-driven clinical decision-making that combines subjective clinical assessment of symptoms and preferences with biological measures such as genetics, biomarkers, imaging, and integrative physiology to derive quantitative risk scores and decision support. In contrast, psychiatry has just begun to scratch the surface of measurement-based care with validated clinical questionnaires. An opportunity exists to improve modern psychiatric care with novel data streams from digital sensors combined with clinical observation and subjective self-report. The prospect of integrating this complex information with modern computational and analytical methods could advance the field, both in research and clinical practice. Here we discuss this possibility and propose some key priorities to enable these innovations toward improving clinical outcomes in the future.

Activation of the Listeria monocytogenes Virulence Program by a Reducing Environment.

Upon entry into the host cell cytosol, the facultative intracellular pathogen Listeria monocytogenes coordinates the expression of numerous essential virulence factors by allosteric binding of glutathione (GSH) to the Crp-Fnr family transcriptional regulator PrfA. Here, we report that robust virulence gene expression can be recapitulated by growing bacteria in a synthetic medium containing GSH or other chemical reducing agents. Bacteria grown under these conditions were 45-fold more virulent in an acute murine infection model and conferred greater immunity to a subsequent lethal challenge than bacteria grown in conventional media. During cultivation in vitro, PrfA activation was completely dependent on the intracellular levels of GSH, as a glutathione synthase mutant (ΔgshF) was activated by exogenous GSH but not reducing agents. PrfA activation was repressed in a synthetic medium supplemented with oligopeptides, but the repression was relieved by stimulation of the stringent response. These data suggest that cytosolic L. monocytogenes interprets a combination of metabolic and redox cues as a signal to initiate robust virulence gene expression in vivoIMPORTANCE Intracellular pathogens are responsible for much of the worldwide morbidity and mortality from infectious diseases. These pathogens have evolved various strategies to proliferate within individual cells of the host and avoid the host immune response. Through cellular invasion or the use of specialized secretion machinery, all intracellular pathogens must access the host cell cytosol to establish their replicative niches. Determining how these pathogens sense and respond to the intracellular compartment to establish a successful infection is critical to our basic understanding of the pathogenesis of each organism and for the rational design of therapeutic interventions. Listeria monocytogenes is a model intracellular pathogen with robust in vitro and in vivo infection models. Studies of the host-sensing and downstream signaling mechanisms evolved by L. monocytogenes often describe themes of pathogenesis that are broadly applicable to less tractable pathogens. Here, we describe how bacteria use external redox states as a cue to activate virulence.

c-di-AMP modulates Listeria monocytogenes central metabolism to regulate growth, antibiotic resistance and osmoregulation.

Cyclic diadenosine monophosphate (c-di-AMP) is a conserved nucleotide second messenger critical for bacterial growth and resistance to cell wall-active antibiotics. In Listeria monocytogenes, the sole diadenylate cyclase, DacA, is essential in rich, but not synthetic media and ΔdacA mutants are highly sensitive to the ß-lactam antibiotic cefuroxime. In this study, loss of function mutations in the oligopeptide importer (oppABCDF) and glycine betaine importer (gbuABC) allowed ΔdacA mutants to grow in rich medium. Since oligopeptides were sufficient to inhibit growth of the ΔdacA mutant we hypothesized that oligopeptides act as osmolytes, similar to glycine betaine, to disrupt intracellular osmotic pressure. Supplementation with salt stabilized the ΔdacA mutant in rich medium and restored cefuroxime resistance. Additional suppressor mutations in the acetyl-CoA binding site of pyruvate carboxylase (PycA) rescued cefuroxime resistance and resulted in a 100-fold increase in virulence of the ΔdacA mutant. PycA is inhibited by c-di-AMP and these mutations prompted us to examine the role of TCA cycle enzymes. Inactivation of citrate synthase, but not down-stream enzymes suppressed ΔdacA phenotypes. These data suggested that c-di-AMP modulates central metabolism at the pyruvate node to moderate citrate production and indeed, the ΔdacA mutant accumulated six times the concentration of citrate present in wild-type bacteria.

Endoscopic Ultrasound-Guided Treatment of Pancreaticocutaneous Fistulas.

Pancreaticocutaneous fistulas (PCFs) may be refractory to medical therapy or endoscopic retrograde cholangopancreaticography. Four patients underwent endoscopic ultrasound-guided management of refractory PCFs, which were internalized by endoscopic ultrasound-guided transmural puncture of the pancreatic duct (n = 2), fistula tract (n = 1), or both (n = 1), with placement of transmural stents providing internal drainage to the stomach (n = 3) or duodenum (n = 1). Drainage from PCFs ceased in all patients, and all percutaneous drains were removed; internal stents were left in place indefinitely. Endoscopic ultrasound-guided interventions may successfully treat PCFs, allowing removal of percutaneous drains, and are an attractive alternative for patients who might otherwise require pancreatic resection.

Kaposi's sarcoma-associated herpesvirus ORF45 mediates transcriptional activation of the HIV-1 long terminal repeat via RSK2.

UNLABELLED: Robust activation of human immunodeficiency virus type 1 (HIV-1) gene expression occurs upon superinfection with Kaposi's sarcoma-associated herpesvirus (KSHV), a common AIDS-associated pathogen. Though the mechanisms underlying this phenotype remain unknown, several KSHV-encoded factors have been reported to stimulate HIV-1 long terminal repeat (LTR) activity. Here, we systematically evaluated the ability of KSHV tegument proteins to modulate the activation of an integrated HIV-1 LTR and revealed that the most potent individual activator is ORF45. ORF45 directs an increase in RNA polymerase II recruitment to the HIV-1 LTR, leading to enhanced transcriptional output. ORF45 is a robust activator of the p90 ribosomal S6 kinases (RSK), and we found that this activity is necessary but not sufficient to increase transcription from the LTR. Of the three widely expressed RSK isoforms, RSK2 appears to be selectively involved in LTR stimulation by both KSHV ORF45 and HIV-1 Tat. However, constitutively active RSK2 is unable to stimulate the LTR, suggesting that ORF45 may preferentially direct this kinase to a specific set of targets. Collectively, our findings reveal a novel transcriptional activation function for KSHV ORF45 and highlight the importance of RSK2 in shaping the transcriptional environment during infection. IMPORTANCE: Kaposi's sarcoma-associated herpesvirus (KSHV) is a prominent AIDS-associated pathogen. Previous studies have shown that infection of cells containing human immunodeficiency virus type 1 (HIV-1) with KSHV leads to potent stimulation of HIV-1 gene expression by activating the HIV-1 promoter, termed the long terminal repeat (LTR). Here, we compared the abilities of various KSHV proteins to activate gene expression from the HIV-1 LTR and found that KSHV ORF45 is the most potent activator. ORF45 is known to induce cell signaling through ribosomal S6 kinase (RSK) and enhance protein translation. However, we revealed that the activation of a specific isoform of RSK by ORF45 also leads to increased mRNA synthesis from the LTR by the host RNA polymerase. Collectively, our findings provide new insight into the interviral interactions between KSHV and HIV that may ultimately impact disease.

Bier block regional anesthesia and casting for forearm fractures: safety in the pediatric emergency department setting.

BACKGROUND: Bier block regional anesthesia was first described in 1908; however, it is uncommonly used for fears of cardiac and neurological complications. Although recent studies have documented safe usage in an adult population, no study to date has investigated its use in a pediatric setting. In addition, most emergency departments feel that splint placement is safer than casting after acute forearm fracture reduction in the pediatric population. However, to our knowledge there is no such study that documents the complication rates associated with immediate casting. The goal of this study was to assess the safety and efficacy of Bier block regional anesthesia and immediate cast application after closed reduction of pediatric forearm fractures. METHODS: A retrospective review was conducted of patients treated for forearm fractures in a 2-year period at a major metropolitan pediatric hospital. Rates of complications and length and costs of the 2 procedures were analyzed. RESULTS: A total of 600 patients were treated with Bier block regional anesthesia and 645 were treated with conscious sedation for displaced fractures of the forearm in the 2-year study period. No complications requiring admission were seen in either group. No patient experienced compartment syndrome or a need for readmission secondary to cast application. 2.2% and 4.3% (P=0.0382) of patients in the Bier block and sedation groups, respectively, needed their cast bivalved secondary to swelling. The average time from initiation of procedural sedation to discharge was 1 hour and 42 minutes, whereas the time to discharge from initiation of Bier block regional anesthesia was 47 minutes (P<0.0001). The average cost for a patient treated with procedural sedation was $6313, whereas the average cost for the Bier block regional anesthesia group was $4956. CONCLUSIONS: Bier block regional anesthesia is a safe, efficient, and cost-effective method of reducing pediatric forearm fractures. Immediate cast application can be used without fear of major complications. LEVELS OF EVIDENCE: Level III--retrospective review.

Putting the pieces together: integrative modeling platform software for structure determination of macromolecular assemblies.

A set of software tools for building and distributing models of macromolecular assemblies uses an integrative structure modeling approach, which casts the building of models as a computational optimization problem where information is encoded into a scoring function used to evaluate candidate models.

Hand and microvascular replantation call availability study: a national real-time survey of level-I and level-II trauma centers.

BACKGROUND: Inconsistent availability of subspecialty hand and microvascular emergency call services could influence patient outcomes and the efficiency of a system dependent on limited resources and timely intervention because declining reimbursements, increased medicolegal risk, lack of confidence in microsurgical skills, and the disruption of elective schedules present a deterrent to call panel participation. This study assessed the availability of hand and microvascular replantation surgery call services at all level-I and level-II trauma centers in the United States. METHODS: Between May and December 2010, all level-I (N = 137) and level-II (N = 153) trauma centers across the U.S. were contacted by telephone. Phone contact was unannounced; responders were invited to participate in our institutional review board-approved anonymous survey regarding hand and microvascular replantation emergency coverage specific to their hospital. RESULTS: Level-I trauma centers: 117 (85%) of 137 participated, and sixty-four (55%) of these had immediate access for hand surgery and microvascular replantation services. Six hospitals provided services for fifteen to thirty-one days per month, and three hospitals supported services for one to fifteen days per month. Ten hospitals indicated inconsistent coverage, which was difficult to estimate, and thirty-four hospitals reported no coverage. Level-II trauma centers: 132 (86%) of 153 participated, and thirty-eight (29%) of these had immediate access for hand surgery and microvascular replantation services. Seven hospitals provided services for fifteen to thirty-one days per month, and three hospitals provided coverage for one to fifteen days per month. Eighty-four hospitals reported no specific coverage protocol. CONCLUSIONS: Inconsistency in the definition and coverage of emergency hand and microvascular replantation services was identified at level-I and level-II trauma centers across the U.S. Many hospitals indicated the presence of subspecialty hand surgery coverage; however, the determination of microvascular replantation resources was not available consistently. The results of our study strengthen previous conclusions about the need for a more defined and coordinated system of emergency microvascular replantation surgery services in order to improve the efficiency of a limited resource and, ultimately, improve patient care.

Compartment syndrome of the upper extremity.

Compartment syndrome involves the sustained elevation of interstitial tissue pressures within an osteofascial envelope to nonphysiologic levels. Tissue injury involves a spectrum from reversible to irreversible damage and, therefore, early recognition and treatment is critical for optimal outcomes. This article reviews the nature of upper extremity compartment syndrome; considers the general classification scheme and potential causes; and discusses the pertinent anatomy, pathophysiology, treatment recommendations, and outcomes for this challenging condition.

Arthroscopic treatment of osteoarthritis of the knee: are there any evidence-based indications?

Despite the lack of consensus guidelines and randomized control trials, the use of arthroscopy for the treatment of osteoarthritis of the knee has increased over the last decade. Techniques used for the arthroscopic treatment of osteoarthritis of the knee include joint lavage, joint débridement, meniscectomy, abrasion arthroplasty, and microfracture. We performed a retrospective, evidence-based review of the current literature on the arthroscopic treatment of osteoarthritis of the knee and provide insight into the study design flaws and difficulties associated with the current research on this controversial topic. Our literature search yielded 18 relevant studies. Of these, one was Level I evidence, five were Level II, six were Level III, and six were Level IV. We found limited evidence-based research to support the use of arthroscopy as a treatment method for osteoarthritis of the knee. Arthroscopic débridement of meniscus tears and knees with low-grade osteoarthritis may have some utility, but it should not be used as a routine treatment for all patients with knee osteoarthritis.

The diagnosis of meniscus tears: the role of MRI and clinical examination.

Magnetic resonance imaging (MRI) and clinical examination are tools commonly used in the diagnosis of meniscus tears. It has been suggested routine MRI before therapeutic arthroscopy for clinically diagnosed meniscus tears will reduce the number and cost of unnecessary invasive procedures. We designed a systematic review of prospective cohort studies comparing MRI and clinical examination to arthroscopy to diagnosis meniscus tears. Thirty-two relevant studies were identified by a literature review. Careful evaluation by an experienced examiner identifies patients with surgically treatable meniscus lesions with equal or better reliability than MRI. MRI is superior when indications for arthroscopy are solely diagnostic. However, the methods by which such a clinician arrives at a conclusion have not been identified. To create an evidence-based algorithm for the diagnosis of a meniscus tear future investigations should prospectively assess the value of commonly used aspects of the patient history and meniscus tests. MRI is useful, but should be reserved for situations in which an experienced clinician requires further information before arriving at a diagnosis. Indications for arthroscopy should be therapeutic, not diagnostic in nature.