Coupling of mRNA Structure Rearrangement to Ribosome Movement during Bypassing of Non-coding Regions.

Nearly half of the ribosomes translating a particular bacteriophage T4 mRNA bypass a region of 50 nt, resuming translation 3' of this gap. How this large-scale, specific hop occurs and what determines whether a ribosome bypasses remain unclear. We apply single-molecule fluorescence with zero-mode waveguides to track individual Escherichia coli ribosomes during translation of T4's gene 60 mRNA. Ribosomes that bypass are characterized by a 10- to 20-fold longer pause in a non-canonical rotated state at the take-off codon. During the pause, mRNA secondary structure rearrangements are coupled to ribosome forward movement, facilitated by nascent peptide interactions that disengage the ribosome anticodon-codon interactions for slippage. Close to the landing site, the ribosome then scans mRNA in search of optimal base-pairing interactions. Our results provide a mechanistic and conformational framework for bypassing, highlighting a non-canonical ribosomal state to allow for mRNA structure refolding to drive large-scale ribosome movements.

'When they do that kind of bad things to me, I feel that they encourage me to be strong and be who I am': How transfeminine people in Samoa, Tonga and the Cook Islands experience discrimination, acceptance and a sense of place.

Data on transfeminine participants from a 2016 Pacific Multi-Country Mapping and Behavioural Study evidence high levels of verbal, physical and sexual abuse, as well as discrimination. In interviews from the same study, accounts of hardship were frequently countered with assertions of happiness and talk of acceptance. This paper analyses these accounts and, in particular, the ways in which interviewees viewed and managed their place in society. Data provide insights into the factors that support transfeminine occupation of a positive place in some contemporary Pacific settings, highlighting negotiation between modern and traditional, and local and global, cultures and values.

Nisin M: a Bioengineered Nisin A Variant That Retains Full Induction Capacity but Has Significantly Reduced Antimicrobial Activity.

Nisin A is a potent antimicrobial with potential as an alternative to traditional antibiotics, and a number of genetically modified variants have been created that target clinically relevant pathogens. In addition to antimicrobial activity, nisin autoregulates its own production via a signal transduction pathway, a property that has been exploited in a protein expression system termed the nisin-controlled gene expression (NICE) system. Although NICE has become one of the most popular protein expression systems, one drawback is that the inducer peptide, nisin A, also has inhibitory activity. It has already been demonstrated that the N-terminal region of nisin A contributes to antimicrobial activity and signal transduction properties; therefore, we conducted bioengineering of nisin at positions Pro9 and Gly10 within ring B to produce a bank of variants that could potentially be used as alternative induction peptides. One variant, designated nisin M, has threonines at positions 9 and 10 and retains induction capacity comparable to that of wild-type nisin A, while most of the antimicrobial activity is abolished. Further analysis confirmed that nisin M produces a mix of peptides as a result of different degrees of dehydration of the two threonines. We show that nisin M exhibits potential as a more suitable alternative to nisin A for the expression of proteins that may be difficult to express or for production of proteins in strains that are sensitive to wild-type nisin. Moreover, it may address the increasing demand by industry for optimization of peptide fermentations to increase yields or production rates.IMPORTANCE This study describes the generation of a nisin variant with superior characteristics for use in the NICE protein expression system. The variant, termed nisin M, retains an induction capacity comparable to that of wild-type nisin A but exhibits significantly reduced antimicrobial activity and can therefore be used at concentrations that are normally toxic to the expression host.

Guidelines for the investigation of chronic diarrhoea in adults: British Society of Gastroenterology, 3rd edition.

Chronic diarrhoea is a common problem, hence clear guidance on investigations is required. This is an updated guideline from 2003 for the investigations of chronic diarrhoea commissioned by the Clinical Services and Standards Committee of the British Society of Gastroenterology (BSG). This document has undergone significant revision in content through input by 13 members of the Guideline Development Group (GDG) representing various institutions. The GRADE system was used to appraise the quality of evidence and grading of recommendations.

Understanding the meaning of youth sexual and reproductive well-being in Fiji.

Well-being is a term commonly used in discussions of sexuality, reproduction and sexual health, yet the meaning of the term is elusive and often disregarded. As an example, the 'well-being' component of sexual and reproductive health and well-being is often not explicitly addressed in research, policy and programme development. The goal of this paper is to explore the meanings of sexual and reproductive well-being among young people in Fiji and their implications. Fourteen focus group discussions with young people aged 15 to 19 years and 40 key informant interviews were held in four Fijian settings. We found both different and shared meanings of sexual and reproductive well-being and suggest areas in which it can be strengthened in Fiji.

Knowledge, beliefs, and decisions of pregnant Australian women concerning donation and storage of umbilical cord blood: a population-based survey.

BACKGROUND: Many women giving birth in Australian hospitals can choose to donate their child's umbilical cord blood to a public cord blood bank or pay to store it privately. We conducted a survey to determine the proportion and characteristics of pregnant women who are aware of umbilical cord blood (UCB) banking and who have considered and decided about this option. The survey also sought to ascertain information sources, knowledge, and beliefs about UCB banking, and the effect of basic information about UCB on decisions. METHODS: Researchers and hospital maternity staff distributed a survey with basic information about UCB banking to 1,873 women of at least 24 weeks' gestation who were attending antenatal classes and hospital clinics in 14 public and private maternity hospitals in New South Wales. RESULTS: Most respondents (70.7%) were aware of UCB banking. Their main information sources were leaflets from hospital clinics, print media, antenatal classes, TV, radio, friends, and relatives. Knowledge about UCB banking was patchy, and respondents overestimated the likelihood their child would need or benefit from UCB. Women who were undecided about UCB banking were younger, less educated, or from ethnic or rural backgrounds. After providing basic information about UCB banking, the proportion of respondents who indicated they had decided whether or not to donate or store UCB more than doubled from 30.0 to 67.7 percent. CONCLUSIONS: Basic information for parents about UCB banking can affect planned decisions about UCB banking. Information should be accurate and balanced, should counter misconceptions, and should target specific groups.

Impact of the Thompson method on breastfeeding exclusivity and duration: Multi-method design.

BACKGROUND: How hospital clinicians facilitate breastfeeding in the first 48-72 h is critical to breastfeeding exclusivity and duration. Mothers who discharge hospital directly breastfeeding are more likely to continue exclusively breastfeeding at 3-months. OBJECTIVE: To assess the impact of facility-wide implementation of a physiological breastfeeding method (the Thompson method) on direct breastfeeding at hospital discharge and exclusive breastfeeding at 3-months of age. DESIGN: Multi-method design using interrupted time series analysis and surveys. SETTING(S): An Australian tertiary maternity hospital. PARTICIPANTS: 13,667 mother-baby pairs (interrupted time series analysis) and 495 postnatal mothers (surveys). METHODS: The Thompson method includes cradle position and hold, alignment of mouth-to-nipple, baby-led connection and seal, maternal fine-tuning for symmetry, and leisurely duration. We used a large pre-post implementation dataset and conducted interrupted time series analysis using a 24-month baseline period (January 2016 - December 2017); and a 15-month post-implementation period (April 2018 - June 2019). We recruited a sub-sample of women to complete surveys at hospital discharge and 3-months postpartum. Surveys were primarily used to measure impact of Thompson method on exclusive breastfeeding at 3-months, compared with a baseline survey conducted in same setting. RESULTS: Following implementation of the Thompson method, the declining trend in direct breastfeeding at hospital discharge was significantly averted by 0.39% each month relative to baseline (95% CI: 0.03% to 0.76%; p = 0.037). While the 3-month exclusive breastfeeding rate in the Thompson group was 3 percentage points higher than the baseline group; this result did not reach statistical significance. However, a subgroup analysis of women who discharged hospital exclusively breastfeeding revealed the relative odds of exclusive breastfeeding at 3-months in the Thompson group was 0.25 (95% CI: 0.17 to 0.38; p < 0.001), significantly better than the baseline group (Z = 3.23, p < 0.01) where the relative odds was only 0.07 (95% CI: 0.03 to 0.19; p < 0.001). CONCLUSIONS: Implementation of the Thompson method for well mother-baby pairs improved direct breastfeeding trends at hospital discharge. For women who discharged hospital exclusively breastfeeding, exposure to the Thompson method reduced the risk of exclusive breastfeeding discontinuation by 3-months. The positive impact of the method was potentially confounded by partial implementation and a parallel rise in birth interventions which undermine breastfeeding. We recommend strategies to strengthen clinician buy-in to the method, and future research using a cluster randomised trial design. TWEETABLE ABSTRACT: Facility-wide implementation of the Thompson method improves direct breastfeeding at hospital discharge and predicts breastfeeding exclusivity at 3-months.

Progress towards triple elimination of mother-to-child transmission of HIV, hepatitis B and syphilis in Pacific Island Countries and Territories: a systematic review.

The diverse geographic, demographic, and societal factors in the Pacific Island Countries and Territories (PICTs) have contributed to unique epidemiological patterns of HIV, syphilis, and hepatitis B. Transmission can be during pregnancy, at the time of birth or via breastfeeding for HIV, and can have long-term adverse outcomes. Given the similarities in prevention of mother-to-child transmission of these infections, coordinated interventions for triple elimination are used. This systematic review has evaluated the peer-reviewed literature, grey literature, and global databases to assess the availability of data to report against elimination targets in the WHO Regional Framework for the Triple Elimination of Mother-to-Child Transmission of HIV, Hepatitis B and Syphilis in Asia and the Pacific 2018-2030. The secondary objective is to report on progress towards these targets. The findings show that none of the PICTs are on track to achieve triple elimination by 2030. Amongst the limited publicly available indicator data, there is suboptimal coverage for most indicators. It is important that there is an increase in availability of and access to antenatal care, testing, and treatment for pregnant women. Increased efforts are needed to collect data on key indicators and integrate reporting into existing systems to avoid extra burden. Funding: Leila Bell was supported by an Australian Government Research Training Program (RTP) Scholarship, Australia. Funding sources had no role in paper design, data collection, data analysis, interpretation, or writing of the paper.

Pharmacogenomic knowledge and awareness among diverse patients treated with angiotensin converting enzyme inhibitors.

We developed novel electronic phenotyping algorithms for the BioMe biobank data, which accurately identified angiotensin converting enzyme inhibitor (ACEi)-induced angioedema cases and controls. A survey was mailed to all 1075 patients and 91 were returned. Over a third reported that prescribing physicians had not discussed with them the concepts of interindividual drug response variability or adverse event risk, and 73% of patients were previously unaware of pharmacogenomics; however, most patients were interested in having pharmacogenomic testing. Moreover, 67% of patients indicated that pharmacogenomic testing would positively influence their medication compliance. In addition to identifying an innovative approach to define biobank cohorts for pharmacogenomic studies, these results indicate that patients are interested in pharmacogenomic testing, which could translate to improved adherence.

Translational bypassing without peptidyl-tRNA anticodon scanning of coding gap mRNA.

Half the ribosomes translating the mRNA for phage T4 gene 60 topoisomerase subunit bypass a 50 nucleotide coding gap between codons 46 and 47. The pairing of codon 46 with its cognate peptidyl-tRNA anticodon dissociates, and following mRNA slippage, peptidyl-tRNA re-pairs to mRNA at a matched triplet 5' adjacent to codon 47, where translation resumes. Here, in studies with gene 60 cassettes, it is shown that the peptidyl-tRNA anticodon does not scan the intervening sequence for potential complementarity. However, certain coding gap mutants allow peptidyl-tRNA to scan sequences in the bypassed segment. A model is proposed in which the coding gap mRNA enters the ribosomal A-site and forms a structure that precludes peptidyl-tRNA scanning of its sequence. Dissipation of this RNA structure, together with the contribution of 16S rRNA anti-Shine-Dalgarno sequence pairing with GAG, facilitates peptidyl-tRNA re-pairing to mRNA.

Umbilical cord blood banking: beyond the public-private divide.

Umbilical cord blood is a source of haematopoietic progenitor cells, which are used to treat a range of malignant, genetic, metabolic and immune disorders. Until recently, cord blood was either collected through donations to publicly funded cord blood banks for use in allogeneic transplantation, or stored in commercial cord blood banks for use in autologous transplantation. The line between public and private cord blood banking is being blurred by the emergence of "hybrid" models that combine aspects of both the public and private systems. The authors describe these hybrid models and argue that their emergence is explained by both market forces and public sector policy They propose that the future of the sector will depend heavily on several key developments that will differentially affect public, private and hybrid banking models.

Religious perspectives on umbilical cord blood banking.

Umbilical cord blood is a valuable source of haematopoietic stem cells. There is little information about whether religious affiliations have any bearing on attitudes to and decisions about its collection, donation and storage. The authors provided information about umbilical cord blood banking to expert commentators from six major world religions (Catholicism, Anglicanism, Islam, Judaism, Hinduism and Buddhism) and asked them to address a specific set of questions in a commentary. The commentaries suggest there is considerable support for umbilical cord blood banking in these religions. Four commentaries provide moral grounds for favouring public donation over private storage. None attach any particular religious significance to the umbilical cord or to the blood within it, nor place restrictions on the ethnicity or religion of donors and recipients. Views on ownership of umbilical cord blood vary. The authors offer a series of general points for those who seek a better understanding of religious perspectives on umbilical cord blood banking.

BET Protein Inhibition Regulates Macrophage Chromatin Accessibility and Microbiota-Dependent Colitis.

Introduction: In colitis, macrophage functionality is altered compared to normal homeostatic conditions. Loss of IL-10 signaling results in an inappropriate chronic inflammatory response to bacterial stimulation. It remains unknown if inhibition of bromodomain and extra-terminal domain (BET) proteins alters usage of DNA regulatory elements responsible for driving inflammatory gene expression. We determined if the BET inhibitor, (+)-JQ1, could suppress inflammatory activation of macrophages in Il10-/- mice. Methods: We performed ATAC-seq and RNA-seq on Il10-/- bone marrow-derived macrophages (BMDMs) cultured in the presence and absence of lipopolysaccharide (LPS) with and without treatment with (+)-JQ1 and evaluated changes in chromatin accessibility and gene expression. Germ-free Il10-/- mice were treated with (+)-JQ1, colonized with fecal slurries and underwent histological and molecular evaluation 14-days post colonization. Results: Treatment with (+)-JQ1 suppressed LPS-induced changes in chromatin at distal regulatory elements associated with inflammatory genes, particularly in regions that contain motifs for AP-1 and IRF transcription factors. This resulted in attenuation of inflammatory gene expression. Treatment with (+)-JQ1 in vivo resulted in a mild reduction in colitis severity as compared with vehicle-treated mice. Conclusion: We identified the mechanism of action associated with a new class of compounds that may mitigate aberrant macrophage responses to bacteria in colitis.

YgdE is the 2'-O-ribose methyltransferase RlmM specific for nucleotide C2498 in bacterial 23S rRNA.

The rRNAs of Escherichia coli contain four 2'-O-methylated nucleotides. Similar to other bacterial species and in contrast with Archaea and Eukaryota, the E. coli rRNA modifications are catalysed by specific methyltransferases that find their nucleotide targets without being guided by small complementary RNAs. We show here that the ygdE gene encodes the methyltransferase that catalyses 2'-O-methylation at nucleotide C2498 in the peptidyl transferase loop of E. coli 23S rRNA. Analyses of rRNAs using MALDI mass spectrometry showed that inactivation of the ygdE gene leads to loss of methylation at nucleotide C2498. The loss of ygdE function causes a slight reduction in bacterial fitness. Methylation at C2498 was restored by complementing the knock-out strain with a recombinant copy of ygdE. The recombinant YgdE methyltransferase modifies C2498 in naked 23S rRNA, but not in assembled 50S subunits or ribosomes. Nucleotide C2498 is situated within a highly conserved and heavily modified rRNA sequence, and YgdE's activity is influenced by other modification enzymes that target this region. Phylogenetically, YgdE is placed in the cluster of orthologous groups COG2933 together with S-adenosylmethionine-dependent, Rossmann-fold methyltransferases such as the archaeal and eukaryotic RNA-guided fibrillarins. The ygdE gene has been redesignated rlmM for rRNA large subunit methyltransferase M.

How well does a shortened time interval characterize results of a full ambulatory blood pressure monitoring session?

Ambulatory blood pressure monitoring (ABPM) is useful in evaluating cardiovascular risk but requires significant time. The authors examined how closely shortened time intervals correlate with the systolic blood pressure (BP) determined from a full 24-hour ABPM session in 1004 ABPM recordings. After excluding the first hour, Pearson correlations performed for the mean systolic BP of the subsequent 3-, 5-, and 7-hour periods (4, 6, and 8 hours total) with the entire, and remainder of the session, demonstrated greatest improvement in correlation when the session is increased from 4 to 6 hours. Bland-Altman analysis of the 6-hour time period revealed a mean difference of 5.41 mm Hg compared with the full session mean. The authors conclude that 6-hour ABPM can approximate the overall mean BP obtained from full 24-hour ABPM. However, shortened sessions do not characterize the influence of circadian variation on the 24-hour mean BP and may overestimate the 24-hour BP levels.

Bedside Assessment of the Necessity of Daily Lab Testing for Patients Nearing Discharge.

As part of the Choosing Wisely® campaign, the Society of Hospital Medicine recommends against performing "repetitive complete blood count chemistry testing in the face of clinical and lab stability." With this recommendation as a framework, we targeted 2 hospitalist-run inpatient medicine units that employed bedside, scripted, interdisciplinary rounds. Our multifaceted intervention included prompting the hospitalist to identify clinically stable patients for next-day discharge and to discontinue labs when appropriate. It was coupled with the education of the clinicians and a regular data review for the hospitalists and unit staff. Among 2877 discharges included in a 1-year period, there was a significantly decreasing trend after the intervention in the percentage of patients getting labs in the 24, 48, and 72 hours before discharge (-1.87%, -1.47%, and -0.74% decrease per month, respectively; P < 0.05). Our structured, multifaceted approach effectively reduced daily lab testing in the 24 to 48 hours prior to discharge.

Predictors of breastfeeding exclusivity and duration in a hospital without Baby Friendly Hospital Initiative accreditation: A prospective cohort study.

AIM: The aim of this study was to investigate the maternity care factors associated with exclusive breastfeeding duration at three months and six months postpartum in a setting without BFHI accrediation. METHODS: A prospective cohort design. Participants from one tertiary maternity hospital were eligible if they intended to exclusively breastfeed, had birthed a live, term baby; were breastfeeding at recruitment; were rooming-in with their baby; were healthy and well; and understood English. Participants completed an infant feeding survey using 24-h recall questions at three time-points. Data were analysed using descriptive statistics, bivariate analysis and regression modelling. FINDINGS: We recruited 424 participants of whom 84% (n=355) responded to the survey at 3-months and 79% (n=335) at 6-months. Women who avoided exposure to intrapartum opioid analgesia (e.g. intramuscular, intraveous or epidural) were more likely to be exclusively breastfeeding at 3-months postpartum (adjusted odds ratio (aOR) 2.09, 95% confidence interval (CI) 1.15-3.80, probability value (p) 0.016). The only other modifiable predictor of exclusive breastfeeding at 3-months was non-exposure to artificial formula on the postnatal ward (aOR 2.44, 95% CI 1.43-4.18, p<0.001). At 6-months postpartum, the rate of exclusive breastfeeding had reduced to 5% (n=16) which rendered regression modelling untenable. DISCUSSION: Strategies to decrease exposure to opioid analgesia in birth settings and the use of infant formula on the postnatal ward may improve exclusive breastfeeding at three months. CONCLUSION: Results suggest that both intrapartum and postpartum maternity care practices can predict long-term breastfeeding success.

Colonic epithelial miR-31 associates with the development of Crohn's phenotypes.

BACKGROUND: Crohn's disease (CD) is highly heterogeneous, due in large part to variability in cellular processes that underlie the natural history of CD, thereby confounding effective therapy. There is a critical need to advance understanding of the cellular mechanisms that drive CD heterogeneity. METHODS: We performed small RNA sequencing of adult colon tissue from CD and NIBD controls. Colonic epithelial cells and immune cells were isolated from colonic tissues, and microRNA-31 (miR-31) expression was measured. miR-31 expression was measured in colonoid cultures generated from controls and patients with CD. We performed small RNA-sequencing of formalin-fixed paraffin-embedded colon and ileum biopsies from treatment-naive pediatric patients with CD and controls and collected data on disease features and outcomes. RESULTS: Small RNA-sequencing and microRNA profiling in the colon revealed 2 distinct molecular subtypes, each with different clinical associations. Notably, we found that miR-31 expression was a driver of these 2 subtypes and, further, that miR-31 expression was particularly pronounced in epithelial cells. Colonoids revealed that miR-31 expression differences are preserved in this ex vivo system. In adult patients, low colonic miR-31 expression levels at the time of surgery were associated with worse disease outcome as measured by need for an end ileostomy and recurrence of disease in the neoterminal ileum. In pediatric patients, lower miR-31 expression at the time of diagnosis was associated with future development of fibrostenotic ileal CD requiring surgeryCONCLUSIONS. These findings represent an important step forward in designing more effective clinical trials and developing personalized CD therapies. FUNDING: This work was supported by CCF Career Development Award (SZS), R01-ES024983 from NIEHS (SZS and TSF), 1R01DK104828-01A1 from NIDDK (SZS and TSF), P01-DK094779-01A1 from NIDDK (SZS), P30-DK034987 from NIDDK (SZS), 1-16-ACE-47 ADA Pathway Award (PS), UNC Nutrition Obesity Research Center Pilot & Feasibility Grant P30DK056350 (PS), CCF PRO-KIIDS NETWORK (SZS and PS), UNC CGIBD T32 Training Grant from NIDDK (JBB), T32 Training Grant (5T32GM007092-42) from NIGMS (MH), and SHARE from the Helmsley Trust (SZS). The UNC Translational Pathology Laboratory is supported, in part, by grants from the National Cancer Institute (3P30CA016086) and the UNC University Cancer Research Fund (UCRF) (PS).

How bad is bile acid diarrhoea: an online survey of patient-reported symptoms and outcomes.

OBJECTIVES: Bile acid diarrhoea (BAD) is an underdiagnosed condition producing diarrhoea, urgency and fear of faecal incontinence. How patients experience these symptoms has not previously been studied. Bile Acid Malabsorption (BAM) Support UK was established in 2015 as a national charity with objectives including to provide details regarding how BAD affects patients, to improve earlier recognition and clinical management. DESIGN SETTING AND MAIN OUTCOME: A questionnaire was collected anonymously by BAM Support UK and the Bile Salt Malabsorption Facebook group over 4 weeks at the end of 2015. It comprised 56 questions and aimed to inform patients and clinicians about how BAD affects the respondents. RESULTS: The first 100 responses were analysed. 91% of the respondents reported a diagnosis of BAD. 58% of total respondents diagnosed following a Selenium-homocholic acid taurine scan, 69% were diagnosed by a gastroenterologist, with type 2 and 3 BAD comprising 38% and 37%, respectively, of total respondents. Symptoms had been experienced for more than 5 years before diagnosis in 44% of respondents. Following treatment, usually with bile acid sequestrants, 60% of participants reported improvement of diarrhoea and most reported their mental health has been positively impacted. Just over half of the cohort felt as though their symptoms had been dismissed during clinical consultations and 28% felt their GPs were unaware of BAD. CONCLUSIONS: BAD requires more recognition by clinicians to address the current delays in diagnosis. Treatment improves physical and mental symptoms in the majority of participants.

Strengthening value-based medication management in a free clinic for the uninsured: Quality interventions aimed at reducing costs and enhancing adherence.

Skyrocketing costs of prescription medications in the USA pose a significant threat to the financial viability of safety net clinics that opt to supply medications at low to no out-of-pocket costs to patients. At the East Harlem Health Outreach Partnership clinic of the Icahn School of Medicine at Mount Sinai, a physician-directed student-run comprehensive primary care clinic for uninsured adults of East Harlem, expenditures on pharmaceuticals represent nearly two-thirds of annual costs. The practice of minimising costs while maintaining quality, referred to as high-value care, represents a critical cost-saving opportunity for safety net clinics as well as for more economical healthcare in general. In this paper, we discuss a series of quality improvement initiatives aimed at reducing pharmacy-related expenditures through two distinct yet related mechanisms: (A) promoting value-conscious prescribing by providers and (B) improving patient adherence to medication regimens. Interventions aimed at promoting value-conscious prescribing behaviour included blacklisting a costly medication on our clinic's formulary and adding a decision tree in our mobile clinician reference application to promote value-conscious prescribing. Interventions targeted to improving patient adherence involved an automated text messaging system with English and Spanish refill reminders to encourage timely pick-up of medication refills. As a result of these processes, the free clinic experienced a 7.3%, or $3768, reduction in annual pharmacy costs. Additionally, medication adherence in patients with diabetes on oral antihyperglycaemic medications increased from 55% to 67%. Simultaneous patient-based and provider-based interventions may be broadly applicable to addressing rising pharmacy costs in healthcare across the USA.