High-Resolution Melting (HRM) analysis of DNA methylation using semiconductor chip-based digital PCR.

BACKGROUND: Digital PCR (dPCR) technology allows absolute quantification and detection of disease-associated rare variants, and thus the use of dPCR technology has been increasing in clinical research and diagnostics. The high-resolution melting curve analysis (HRM) of qPCR is widely used to distinguish true positives from false positives and detect rare variants. In particular, qPCR-HRM is commonly used for methylation assessment in research and diagnostics due to its simplicity and high reproducibility. Most dPCR instruments have limited fluorescence channels available and separate heating and imaging systems. Therefore, it is difficult to perform HRM analysis using dPCR instruments. OBJECTIVE: A new digital real-time PCR instrument (LOAA) has been recently developed to integrate partitioning, thermocycling, and imaging in a single dPCR instrument. In addition, a new technique to perform HRM analysis is utilized in LOAA. The aim of the present study is to evaluate the efficiency and accuracy of LOAA dPCR on HRM analysis for the detection of methylation. METHODS: In this study, comprehensive comparison with Bio-Rad qRT-PCR and droplet-based dPCR equipment was performed to verify the HRM analysis-based methylation detection efficiency of the LOAA digital PCR equipment. Here, sodium bisulfite modification method was applied to detect methylated DNA sequences by each PCR method. RESULTS: Melting curve analysis detected four different Tm values using LOAA and qPCR, and found that LOAA, unlike qPCR, successfully distinguished between different Tm values when the Tm values were very similar. In addition, melting temperatures increased by each methylation were about 0.5℃ for qPCR and about 0.2 ~ 0.6℃ for LOAA. The melting temperature analyses of methylated and unmethylated DNA samples were conducted using LOAA dPCR with TaqMan probes and EvaGreen, and the result found that Tm values of methylated DNA samples are higher than those of unmethylated DNA samples. CONCLUSION: The present study shows that LOAA dPCR could detect different melting temperatures according to methylation status of target sequences, indicating that LOAA dPCR would be useful for diagnostic applications that require the accurate quantification and assessment of DNA methylation.

How predictive is peer review for gauging impact? The association between reviewer rating scores, publication status, and article impact measured by citations in a pain subspecialty journal.

BACKGROUND: Peer review represents a cornerstone of the scientific process, yet few studies have evaluated its association with scientific impact. The objective of this study is to assess the association of peer review scores with measures of impact for manuscripts submitted and ultimately published. METHODS: 3173 manuscripts submitted to Regional Anesthesia & Pain Medicine (RAPM) between August 2018 and October 2021 were analyzed, with those containing an abstract included. Articles were categorized by topic, type, acceptance status, author demographics and open-access status. Articles were scored based on means for the initial peer review where each reviewer's recommendation was assigned a number: 5 for 'accept', 3 for 'minor revision', 2 for 'major revision' and 0 for 'reject'. Articles were further classified by whether any reviewers recommended 'reject'. Rejected articles were analyzed to determine whether they were subsequently published in an indexed journal, and their citations were compared with those of accepted articles when the impact factor was <1.4 points lower than RAPM's 5.1 impact factor. The main outcome measure was the number of Clarivate citations within 2 years from publication. Secondary outcome measures were Google Scholar citations within 2 years and Altmetric score. RESULTS: 422 articles met inclusion criteria for analysis. There was no significant correlation between the number of Clarivate 2-year review citations and reviewer rating score (r=0.038, p=0.47), Google Scholar citations (r=0.053, p=0.31) or Altmetric score (p=0.38). There was no significant difference in 2-year Clarivate citations between accepted (median (IQR) 5 (2-10)) and rejected manuscripts published in journals with impact factors >3.7 (median 5 (2-7); p=0.39). Altmetric score was significantly higher for RAPM-published papers compared with RAPM-rejected ones (median 10 (5-17) vs 1 (0-2); p<0.001). CONCLUSIONS: Peer review rating scores were not associated with citations, though the impact of peer review on quality and association with other metrics remains unclear.

Thoracic sympathetic ganglion blocks: real-world outcomes in 207 chronic pain patients.

BACKGROUND: Thoracic sympathetic ganglion block (TSGB) is a procedure to manage sympathetically maintained upper extremity pain (sympathetically maintained pain). To date, only a few studies have evaluated the clinical effectiveness of TSGB in pain medicine. This study investigated (1) the relationship between technical success of TSGB and pain reduction in patients with chronic upper extremity pain and (2) relevant clinical factors for a positive TSGB outcome. METHODS: We retrospectively reviewed medical data in 232 patients who received TSGB from 2004 to 2020. Technical success and a positive outcome of TSGB were defined as a temperature increase of ≥1.5°C at 20 min and a pain reduction with ≥2 points on the 11-point Numerical Rating Scale at 2 weeks post-TSGB, respectively. Correlations were assessed using correlation coefficients (R), and multivariable regression model was used to identify factors relevant to TSGB outcomes. RESULTS: 207 patients were ultimately analyzed; among them, 115 (55.5%) patients positively responded to TSGB, and 139 (67.1%) achieved technical success after TSGB. No significant relationship existed between the pain reduction and the temperature increase after TSGB (R=0.013, p=0.855). Comorbid diabetes (OR 4.200) and adjuvant intake (OR 3.451) were positively associated, and psychiatric comorbidity (OR 0.327) and pain duration (OR 0.973) were negatively associated with TSGB outcome. CONCLUSIONS: We found no significant association between the temperature increase and pain reduction after TSGB. Further studies are warranted to identify significant factors associated with TSGB outcomes in patients with complex regional pain syndrome and neuropathic pain diseases.

Myth and Truth in Opioid Consumption with Intrathecal Morphine Pump Implantation in Chronic Pain: A Retrospective Cohort Study with Claims Database in South Korea.

OBJECTIVE: To investigate the opioid consumption and the healthcare resource utilization in patients with the intrathecal drug delivery system (IDDS) therapy and the comprehensive medical management (CMM) alone. DESIGN: A retrospective cohort study with a customized claims database. SETTING: In a university-based hospital. SUBJECTS: Patients with complex regional pain syndrome, post-laminectomy syndrome, and fibromyalgia. METHODS: Using propensity score matching (1:3), we selected patients with morphine infusion through IDDS (IDDS group) and CMM alone (CMM group). The primary endpoints were comparisons of average morphine equivalents daily dosages (MEDD, mg/day) for 6 and 12 months from an index date. The number of emergency room (ER) visits and hospitalizations and the total medical expenditures were compared as secondary outcomes. RESULTS: In total, 82 patients (N = 23 in the IDDS group and N = 59 in the CMM group) were analyzed. Although a 6-month average MEDD did not reach statistical significance, a 12-month average MEDD was significantly decreased in the IDDS group compared to the CMM group (53.2 ± 46.3 vs 123.9 ± 176.4, respectively; P = 0.008). ER visits were more frequent in the IDDS group than the CMM group at baseline (5.4 vs 0.5, respectively; P = .002), which was maintained for 12 months (P < 0.001). Otherwise, the number of hospitalization and the medical expenditures for pain management were not different between the groups for 12 months. CONCLUSIONS: The combined IDDS therapy had some benefits in reducing opioid consumption for 1-year follow-up compared to the CMM alone in chronic noncancer pain patients.

Eptinezumab-jjmr, a humanized monoclonal specific to Calcitonin Gene Related Peptide, for the preventive treatment of migraine in adults.

Purpose of Review: Migraines are prevalent and cause significant morbidity, decline in quality of life and healthcare costs universally. Treatment options are varied, but efficacy is limited. This review centers on Eptinezumab-jjmr, a humanized monoclonal specific to CGRP for the prevention of migraines in adults. Herein presented are the science and mechanism of action, indication and clinical evidence for use. Recent Findings: Migraines are severe, recurrent headaches, which are either episodic or chronic in nature. The pain is severe, often accompanied by co-morbid symptoms, such as photophobia, phonophobia, nausea and emesis, and is limiting in nature. It is a prevalent disorder that causes significant, worldwide disability, morbidity, suffering, and costs.The pathophysiology of migraines is actively studied, though recent research points to an initiating event causing migraine generation, that is then propagated by other brain regions, a significant one being the trigeminocervical complex. This is driven by biochemical transmitters, chiefly CGRP. This discovery led to the development of CGRP-targeting drugs, including gepants (small molecular antagonists) and anti-CGRP antibodies, such as Eptinezumab-jjmr.Traditional therapy includes preventative and abortive treatment; however, adherence with preventative treatment has been historically poor, and certain types of abortive therapy carry risks and side effects that preclude them from a large patient population. Moreover, traditional therapy often falls short in migraine therapy. CGRP antagonist, including Eptinezumab, aims to cover the gaps in migraine therapy. We present here evidence to support the safe and effective use of Eptinezumab for the prevention of migraines. Summary: Migraines are a prevalent primary headache disorder causing significant morbidity worldwide. Traditional abortive and preventative treatments fall short for many patients. Eptinezumab is part of new generation of CGRP-targeting medications and has shown significant evidence to support its use for the prevention of migraines. Further research is required to properly compare eptinezumab with existing pharmacotherapy and update guidelines on the appropriate combinations of therapies that are not available and the correct patient selection for each.

Strain level and comprehensive microbiome analysis in inflammatory bowel disease via multi-technology meta-analysis identifies key bacterial influencers of disease.

Objective: Inflammatory bowel disease (IBD) is a heterogenous disease in which the microbiome has been shown to play an important role. However, the precise homeostatic or pathological functions played by bacteria remain unclear. Most published studies report taxa-disease associations based on single-technology analysis of a single cohort, potentially biasing results to one clinical protocol, cohort, and molecular analysis technology. To begin to address this key question, precise identification of the bacteria implicated in IBD across cohorts is necessary. Methods: We sought to take advantage of the numerous and diverse studies characterizing the microbiome in IBD to develop a multi-technology meta-analysis (MTMA) as a platform for aggregation of independently generated datasets, irrespective of DNA-profiling technique, in order to uncover the consistent microbial modulators of disease. We report the largest strain-level survey of IBD, integrating microbiome profiles from 3,407 samples from 21 datasets spanning 15 cohorts, three of which are presented for the first time in the current study, characterized using three DNA-profiling technologies, mapping all nucleotide data against known, culturable strain reference data. Results: We identify several novel IBD associations with culturable strains that have so far remained elusive, including two genome-sequenced but uncharacterized Lachnospiraceae strains consistently decreased in both the gut luminal and mucosal contents of patients with IBD, and demonstrate that these strains are correlated with inflammation-related pathways that are known mechanisms targeted for treatment. Furthermore, comparative MTMA at the species versus strain level reveals that not all significant strain associations resulted in a corresponding species-level significance and conversely significant species associations are not always re-captured at the strain level. Conclusion: We propose MTMA for uncovering experimentally testable strain-disease associations that, as demonstrated here, are beneficial in discovering mechanisms underpinning microbiome impact on disease or novel targets for therapeutic interventions.

Opicapone, a Novel Catechol-O-methyl Transferase Inhibitor, for Treatment of Parkinson's Disease "Off" Episodes.

Parkinson's Disease (PD) is a common neurodegenerative disorder and the leading cause of disability. It causes significant morbidity and disability through a plethora of symptoms, including movement disorders, sleep disturbances, and cognitive and psychiatric symptoms. The traditional pathogenesis theory of PD involves the loss of dopaminergic neurons in the substantia nigra (SN). Classically, treatment is pursued with an assortment of medications that are directed at overcoming this deficiency with levodopa being central to most treatment plans. Patients taking levodopa tend to experience "off episodes" with decreasing medication levels, causing large fluctuations in their symptoms. These off episodes are disturbing and a source of morbidity for these patients. Opicapone is a novel, peripherally acting Catechol-O-methyl transferase (COMT) inhibitor that is used as adjunctive therapy to carbidopa/levodopa for treatment and prevention of "off episodes." It has been approved for use as an adjunct to levodopa since 2016 in Europe and has recently (April 2020) gained FDA approval for use in the USA. By inhibiting COMT, opicapone slows levodopa metabolism and increases its availability. Several clinical studies demonstrated significant improvement in treatment efficacy and reduction in duration of "off episodes." The main side effect demonstrated was dyskinesia, mostly with the 100mg dose, which is higher than the approved, effective dose of 50mg. Post-marketing surveillance and analysis are required to further elucidate its safety profile and contribute to patient selection. This paper reviews the seminal and latest evidence in the treatment of PD "off episodes" with the novel drug Opicapone, including efficacy, safety, and clinical indications.

Glossopharyngeal Neuralgia: Epidemiology, Risk factors, Pathophysiology, Differential diagnosis, and Treatment Options.

Purpose of Review: This is a comprehensive review of the most recent literature on glossopharyngeal neuralgia (GPN), a relatively rare form of neuropathic facial pain. It covers the epidemiology, risk factors, pathophysiology, and differential diagnosis given that glossopharyngeal neuralgia can often be confused with other facial pain syndromes. Finally, we extensively review recent findings regarding medical or conservative measures, minimally invasive, and surgical options for potentially treating and managing glossopharyngeal neuralgia. Recent Findings: An in-depth analysis of the recent literature indicates that glossopharyngeal neuralgia is not only rare but its etiology and pathophysiology are complex and are often secondary to other disease processes. Regardless, current management options are shown to be effective in controlling pain. Conservatively, first-line management of GPN is carbamazepine, but gabapentin and eslicarbazepine acetate are suitable alternatives. In terms of current minimally invasive pain management techniques, pulsed radiofrequency ablation, nerve blocks, or percutaneous radiofrequency thermocoagulation are effective. Finally, surgical management involves microvascular decompression and rhizotomy. Summary: While there are currently many viable options for addressing glossopharyngeal neuralgia pain ranging from conservative to surgical management, the complex nature of GPN etiology, pathophysiology, and involved anatomical structures prompts further research for more effective ways to treat the disease.

Can the Rhomboid Major Muscle Be Used to Identify the Thoracic Spinal Segment on Ultrasonography? A Prospective Observational Study.

OBJECTIVE: We investigated the thoracic segment corresponding to the inferior margin of the rhomboid major muscle (RMM) using ultrasound (US) to evaluate its potential as a reliable anatomic landmark for segment identification. DESIGN: A prospective observational study. SETTING: An operating room. SUBJECTS: Patients who underwent procedures around the thoracic spine. METHODS: Four hundred segments corresponding to the RMM's inferior margin were identified through the use of paravertebral sagittal US and confirmed by fluoroscopy in 100 participants in the prone position with upward and downward shoulder rotation, comprising four datasets (up-right, up-left, down-right, and down-left). The US identification of the RMM's inferior margin was dichotomously scored (clear vs ambiguous). Each dataset was divided into two groups (dominant segment group vs remaining segments group), which were compared. Factors relevant to the dominant segment associated with the RMM's inferior border were determined through univariable analyses. RESULTS: The T6 segment was observed most commonly (59.5%) along the RMM's inferior border on paravertebral sagittal US acquired in the prone position, followed by T5 (25.0%), T7 (12.8%), and T4 (2.7%). The segments corresponding to the RMM remained unchanged by shoulder posture in most participants (n = 74, 74%). The RMM's inferior border was clearly distinguishable in 330 cases (82.5%). When the RMM's inferior border was clearly identified, the corresponding segment was likely to match T6 in all datasets, with odds ratios ranging from 3.24 to 6.2. CONCLUSIONS: The RMM's inferior border over the transverse process corresponded to T6 most frequently on paravertebral sagittal US, and its deep fascia was clearly visible in most cases.

Not FIT for Use: Fecal Immunochemical Testing in the Inpatient and Emergency Settings.

BACKGROUND: Fecal immunochemical testing (FIT) is widely used for colorectal cancer screening, its only indication. Its effect on clinical decision-making beyond screening is unknown. We studied the use of FIT in emergency and inpatient settings and its impact on patient care. METHODS: Using electronic medical records, we reviewed all non-ambulatory FITs performed from November 2017 to October 2019 at a tertiary care community hospital. We collected data on demographics, indications, gastroenterology consultations, and endoscopic procedures. Multivariate logistic regression was performed to determine the effect of FIT on gastroenterology consultation and endoscopy. RESULTS: We identified 550 patients with at least 1 FIT test. Only 3 FITs (0.5%) were performed for colorectal cancer screening. FITs were primarily ordered from the emergency department (45.3%) or inpatient hospital floor (42.2%). Anemia (44.0%), followed by gastrointestinal bleeding (40.9%), were the most common indications. FIT was positive in 253 patients (46.0%), and gastroenterology consultation was obtained for 47.4% (n = 120), compared with 14.5% (n = 43) of the 297 FIT-negative patients (odds ratio 3.28; 95% confidence interval, 2.23-4.82, P < .0001). A potential bleeding source was identified in 80% of patients with reported or witnessed overt gastrointestinal bleeding, a similar proportion (80.7%; P = .92) to patients who were FIT positive with overt gastrointestinal bleeding. Multivariate analysis showed that melena, hematemesis, and a positive FIT were associated with gastroenterology consultation (all P < .05), while only melena (odds ratio 3.34; 95% confidence interval, 1.48-7.54) was associated with endoscopy. CONCLUSIONS: Nearly all emergency department and inpatient FIT use was inappropriate. FIT resulted in more gastroenterology consultation but was not independently associated with inpatient endoscopy.

Suvorexant, a Novel Dual Orexin Receptor Antagonist, for the Management of Insomnia.

Purpose of Review: The present investigation is a comprehensive review regarding the use of Suvorexant for insomnia treatment. It covers the background, pathophysiology, and significance of addressing insomnia, the pharmaceutical details of Suvorexant, and its safety, efficacy, and implications in treating insomnia. We further discuss Suvorexant's role in targeting insomnia with other comorbidities. Recent Findings: Insomnia refers to poor quality and/or quantity of sleep. While there are many existing treatments such as benzodiazepines, melatonin agonists, TCAs, and atypical antipsychotics used to target various receptors involved in normal induction and maintenance of sleep, Suvorexant is an antagonist that specifically targets orexin receptors. Recent clinical studies suggest that Suvorexant is both clinically safe and effective. Quantity and quality of sleep are measured in various ways, yet the consensus points towards Suvorexant's effectiveness in improving sleep time, onset, latency, and quality compared to placebo. In addition to helping improve isolated insomnia, Suvorexant helps improve sleep in patients that have other comorbidities such as obstructive sleep apnea, Alzheimer's disease, dementia, acute stroke, and delirium. While Suvorexant is safe, there are still adverse effects associated with the drug that needs to be considered. The most common adverse effects include dizziness, somnolence, headaches, and cognitive impairment. Summary: Insomnia is a major public health concern that affects many people worldwide and has been linked to many adverse health outcomes. While there are existing treatments that target different receptors and pathways of normal sleep induction and maintenance, Suvorexant is a novel drug that targets dual orexin receptors. Its safety and efficacy, mechanism of action, pharmacokinetic parameters, and relative lack of rebound and withdrawal effects render suvorexant a reliable choice for the treatment of insomnia.

The role of working memory capacity in spatial learning depends on spatial information integration difficulty in the environment.

A substantial amount of research has been conducted to uncover factors underlying the pronounced individual differences in spatial navigation. Spatial working memory capacity (SWM) is shown to be one important factor. In other domains such as reading comprehension, the role of working memory capacity in task performance differences depends on the difficulty of other task demands. In the current study, we investigated whether, similarly, the relationship between SWM and spatial performance was dependent on the difficulty of spatial information integration in the environment. Based on our prior work, spatial information integration difficulty depends on (a) difficulty in observing spatial relationships between locations of interest in the environment and (b) the individual's ability to integrate such relationships. Leveraging virtual reality, we manipulated the difficulty in observing the spatial relationships during learning by changing the visibility of the buildings, and measured individual's self-report sense of direction (SOD) which modulates the ability to integrate such relationships under different degrees of visibility. We consistently found that in the "easy" spatial integration condition (high SOD with high visibility), high SWM did not significantly improve spatial learning. The same pattern was observed in the difficult condition (low SOD with low visibility). On the other hand, high SWM improved spatial learning for medium difficulty (high SOD with low visibility, or vice versa). Together, our results reveal that the role of SWM in spatial learning differences depends on spatial integration difficulty. Our results also have significant applied implications for using virtual reality to target and facilitate spatial learning. (PsycInfo Database Record (c) 2021 APA, all rights reserved).

Colorectal Cancer Screening Decisions in the Opportunistic Setting.

Colorectal cancer incidence and mortality have decreased in the United States in recent decades, largely through opportunistic screening. Although certain organizations have improved internal screening rates by implementing programmatic screening, most of the United States undergoes opportunistic screening. Much effort and resources have been expended comparing screening tests to determine the most effective; however, deeper analysis of the US population has revealed subsets of ethnicities may be grossly underscreened. The most effective screening test remains the test that is completed and adhered to, and a better question may concern the best method of discussing screening.

Sex differences in heart rate responses to occupational stress.

Cardiovascular disease (CVD) remains the leading cause of disease burden globally and chronic stress is associated with increased risk of CVD. Recognition of chronic occupational stressors as a potential contributor to CVD highlights the need to recognize and prevent stress during work. The ubiquity of wearable technology devices to monitor health provides a new opportunity to noninvasively examine the cardiovascular system throughout a work shift. In the current study, we examined changes in heart rate (HR) during a work shift in a retail store setting using 23 healthy female and male subjects that differed in their physical fitness status. Subjects had their HR tracked via an Apple Watch during three typical work shifts. The results demonstrated an increase in HR during a work shift to a level observed during a moderate stressor (resting HR = 83.2 BPM ± 7.8; highest HR mean = 109.1 BPM ± 11.7; p < .0001). Female subjects demonstrated a significantly elevated maximum HR, a larger change in HR, and a larger percent change in HR compared with males (all p < .05). Physical activity status did not influence the observed changes in HR for females or males. Neither the time of day the work shift occurred nor the length of the shift modulated the observed pattern of HR changes. Collectively, our findings demonstrate the potential for wearables in biomedical research and personalized health.

Microbiota-Produced N-Formyl Peptide fMLF Promotes Obesity-Induced Glucose Intolerance.

The composition of the gastrointestinal microbiota and associated metabolites changes dramatically with diet and the development of obesity. Although many correlations have been described, specific mechanistic links between these changes and glucose homeostasis remain to be defined. Here we show that blood and intestinal levels of the microbiota-produced N-formyl peptide, formyl-methionyl-leucyl-phenylalanine, are elevated in high-fat diet-induced obese mice. Genetic or pharmacological inhibition of the N-formyl peptide receptor Fpr1 leads to increased insulin levels and improved glucose tolerance, dependent upon glucagon-like peptide 1. Obese Fpr1 knockout mice also display an altered microbiome, exemplifying the dynamic relationship between host metabolism and microbiota. Overall, we describe a new mechanism by which the gut microbiota can modulate glucose metabolism, providing a potential approach for the treatment of metabolic disease.

Suppression of myopathic lamin mutations by muscle-specific activation of AMPK and modulation of downstream signaling.

Laminopathies are diseases caused by dominant mutations in the human LMNA gene encoding A-type lamins. Lamins are intermediate filaments that line the inner nuclear membrane, provide structural support for the nucleus and regulate gene expression. Drosophila melanogaster models of skeletal muscle laminopathies were developed to investigate the pathological defects caused by mutant lamins and identify potential therapeutic targets. Human disease-causing LMNA mutations were modeled in Drosophila Lamin C (LamC) and expressed in indirect flight muscle (IFM). IFM-specific expression of mutant, but not wild-type LamC, caused held-up wings indicative of myofibrillar defects. Analyses of the muscles revealed cytoplasmic aggregates of nuclear envelope (NE) proteins, nuclear and mitochondrial dysmorphology, myofibrillar disorganization and up-regulation of the autophagy cargo receptor p62. We hypothesized that the cytoplasmic aggregates of NE proteins trigger signaling pathways that alter cellular homeostasis, causing muscle dysfunction. In support of this hypothesis, transcriptomics data from human muscle biopsy tissue revealed misregulation of the AMP-activated protein kinase (AMPK)/4E-binding protein 1 (4E-BP1)/autophagy/proteostatic pathways. Ribosomal protein S6K (S6K) messenger RNA (mRNA) levels were increased and AMPKα and mRNAs encoding downstream targets were decreased in muscles expressing mutant LMNA relative controls. The Drosophila laminopathy models were used to determine if altering the levels of these factors modulated muscle pathology. Muscle-specific over-expression of AMPKα and down-stream targets 4E-BP, Forkhead box transcription factors O (Foxo) and Peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC1α), as well as inhibition of S6K, suppressed the held-up wing phenotype, myofibrillar defects and LamC aggregation. These findings provide novel insights on mutant LMNA-based disease mechanisms and identify potential targets for drug therapy.

Uneven distribution of cobamide biosynthesis and dependence in bacteria predicted by comparative genomics.

The vitamin B12 family of cofactors known as cobamides are essential for a variety of microbial metabolisms. We used comparative genomics of 11,000 bacterial species to analyze the extent and distribution of cobamide production and use across bacteria. We find that 86% of bacteria in this data set have at least one of 15 cobamide-dependent enzyme families, but only 37% are predicted to synthesize cobamides de novo. The distribution of cobamide biosynthesis and use vary at the phylum level. While 57% of Actinobacteria are predicted to biosynthesize cobamides, only 0.6% of Bacteroidetes have the complete pathway, yet 96% of species in this phylum have cobamide-dependent enzymes. The form of cobamide produced by the bacteria could be predicted for 58% of cobamide-producing species, based on the presence of signature lower ligand biosynthesis and attachment genes. Our predictions also revealed that 17% of bacteria have partial biosynthetic pathways, yet have the potential to salvage cobamide precursors. Bacteria with a partial cobamide biosynthesis pathway include those in a newly defined, experimentally verified category of bacteria lacking the first step in the biosynthesis pathway. These predictions highlight the importance of cobamide and cobamide precursor salvaging as examples of nutritional dependencies in bacteria.

TRiC/CCT chaperonins are essential for maintaining myofibril organization, cardiac physiological rhythm, and lifespan.

We recently reported that CCT chaperonin subunits are upregulated in a cardiac-specific manner under time-restricted feeding (TRF) [Gill S et al. (2015) Science 347, 1265-1269], suggesting that TRiC/CCT has a heart-specific function. To understand the CCT chaperonin function in cardiomyocytes, we performed its cardiac-specific knock-down in the Drosophila melanogaster model. This resulted in disorganization of cardiac actin- and myosin-containing myofibrils and severe physiological dysfunction, including restricted heart diameters, elevated cardiac dysrhythmia and compromised cardiac performance. We also noted that cardiac-specific knock-down of CCT chaperonin significantly shortens lifespans. Additionally, disruption of circadian rhythm yields further deterioration of cardiac function of hypomorphic CCT mutants. Our analysis reveals that both the orchestration of protein folding and circadian rhythms mediated by CCT chaperonin are critical for maintaining heart contractility.