Haemolysin coregulated protein is an exported receptor and chaperone of type VI secretion substrates.

Secretion systems require high-fidelity mechanisms to discriminate substrates among the vast cytoplasmic pool of proteins. Factors mediating substrate recognition by the type VI secretion system (T6SS) of Gram-negative bacteria, a widespread pathway that translocates effector proteins into target bacterial cells, have not been defined. We report that haemolysin coregulated protein (Hcp), a ring-shaped hexamer secreted by all characterized T6SSs, binds specifically to cognate effector molecules. Electron microscopy analysis of an Hcp-effector complex from Pseudomonas aeruginosa revealed the effector bound to the inner surface of Hcp. Further studies demonstrated that interaction with the Hcp pore is a general requirement for secretion of diverse effectors encompassing several enzymatic classes. Though previous models depict Hcp as a static conduit, our data indicate it is a chaperone and receptor of substrates. These unique functions of a secreted protein highlight fundamental differences between the export mechanism of T6 and other characterized secretory pathways.

The Value of Community Health Workers in Diabetes Management in Low-Income Populations: A Qualitative Study.

To describe community health workers (CHWs) roles in a diabetes self-management intervention. Retrospective qualitative inductive analysis of open text home visit encounter form from Peer Support for Achieving Independence in Diabetes (Peer AID), a randomized controlled trial in which low-income individuals with poorly controlled diabetes received either CHW home visits or usual care. Following visits, CHWs completed encounter forms documenting the health goal of the visit, the self-management strategies discussed and participant concerns. 634 encounter reports were completed for the 145 intervention participants. CHW notes revealed three main obstacles to optimal disease control: gaps in diabetes knowledge and self-management skills; socioeconomic conditions; and the complexity of the healthcare system. CHWs helped participants overcome these obstacles through extensive, hands-on education, connecting participants to community resources, and assistance navigating the medical system. In addition, the CHWs offered uncomplicated accessibility and availability to their clients. CHWs can be a valuable asset for low-income patients with chronic health conditions who may require more support than what can provided in a typical primary care visit.

Bacterial N-Glycosylation Efficiency Is Dependent on the Structural Context of Target Sequons.

Site selectivity of protein N-linked glycosylation is dependent on many factors, including accessibility of the modification site, amino acid composition of the glycosylation consensus sequence, and cellular localization of target proteins. Previous studies have shown that the bacterial oligosaccharyltransferase, PglB, of Campylobacter jejuni favors acceptor proteins with consensus sequences ((D/E)X1NX2(S/T), where X1,2 ≠ proline) in flexible, solvent-exposed motifs; however, several native glycoproteins are known to harbor consensus sequences within structured regions of the acceptor protein, suggesting that unfolding or partial unfolding is required for efficient N-linked glycosylation in the native environment. To derive insight into these observations, we generated structural homology models of the N-linked glycoproteome of C. jejuni This evaluation highlights the potential diversity of secondary structural conformations of previously identified N-linked glycosylation sequons. Detailed assessment of PglB activity with a structurally characterized acceptor protein, PEB3, demonstrated that this natively folded substrate protein is not efficiently glycosylated in vitro, whereas structural destabilization increases glycosylation efficiency. Furthermore, in vivo glycosylation studies in both glyco-competent Escherichia coli and the native system, C. jejuni, revealed that efficient glycosylation of glycoproteins, AcrA and PEB3, depends on translocation to the periplasmic space via the general secretory pathway. Our studies provide quantitative evidence that many acceptor proteins are likely to be N-linked-glycosylated before complete folding and suggest that PglB activity is coupled to general secretion-mediated translocation to the periplasm. This work extends our understanding of the molecular mechanisms underlying N-linked glycosylation in bacteria.

Structure and regulation of the type VI secretion system.

The type VI secretion system (T6SS) is a complex and widespread gram-negative bacterial export pathway with the capacity to translocate protein effectors into a diversity of target cell types. Current structural models of the T6SS indicate that the apparatus is composed of at least two complexes, a dynamic bacteriophage-like structure and a cell-envelope-spanning membrane-associated assembly. How these complexes interact to promote effector secretion and cell targeting remains a major question in the field. As a contact-dependent pathway with specific cellular targets, the T6SS is subject to tight regulation. Thus, the identification of regulatory elements that control T6S expression continues to shape our understanding of the environmental circumstances relevant to its function. This review discusses recent progress toward characterizing T6S structure and regulation.

Randomized Controlled Trial of a Community Health Worker Self-Management Support Intervention Among Low-Income Adults With Diabetes, Seattle, Washington, 2010-2014.

INTRODUCTION: Community health workers (CHWs) can improve diabetes outcomes; however, questions remain about translating research findings into practical low-intensity models for safety-net providers. We tested the effectiveness of a home-based low-intensity CHW intervention for improving health outcomes among low-income adults with diabetes. METHODS: Low-income patients with glycated hemoglobin A1c (HbA1c) of 8.0% or higher in the 12 months before enrollment from 3 safety-net providers were randomized to a 12-month CHW-delivered diabetes self-management intervention or usual care. CHWs were based at a local health department. The primary outcome was change in HbA1c from baseline enrollment to 12 months; secondary outcomes included blood pressure and lipid levels, quality of life, and health care use. RESULTS: The change in HbA1c in the intervention group (n = 145) (unadjusted mean of 9.09% to 8.58%, change of -0.51) compared with the control group (n = 142) (9.04% to 8.71%, change of -0.33) was not significant (P = .54). In an analysis of participants with poor glycemic control (HbA1c >10%), the intervention group had a 1.23-point greater decrease in HbA1c compared with controls (P = .046). For the entire study population, we found a decrease in reported physician visits (P < .001) and no improvement in health-related quality of life (P = .07) in the intervention group compared with the control group. CONCLUSION: A low-intensity CHW-delivered intervention to support diabetes self-management did not significantly improve HbA1c relative to usual care. Among the subgroup of participants with poor glycemic control (HbA1c >10% at baseline), the intervention was effective.

The Relationship Between Food Insecurity and Depression, Diabetes Distress and Medication Adherence Among Low-Income Patients with Poorly-Controlled Diabetes.

BACKGROUND: Food insecurity- lack of dependable access to adequate food-may play a role in poor diabetes control. OBJECTIVE: We aimed to determine the relationship between food security status and depression, diabetes distress, medication adherence and glycemic control. DESIGN: Secondary analysis of baseline data from Peer Support for Achieving Independence in Diabetes, a randomized controlled trial that enrolled patients from November 2011 to October 2013. PARTICIPANTS: Participants had poorly controlled type 2 diabetes (A1c ≥ 8.0 % on eligibility screen), household income < 250 % of the federal poverty level, were 30-70 years old, and were recruited from a large public hospital, a VA medical center and a community-health center in King County, Washington. MAIN MEASURES: We measured food insecurity determined by the Department of Agriculture's 6-Item Food Security Module. Depression, diabetes distress and medication adherence measured by PHQ-8, Diabetes Distress Scale and Morisky Medication Adherence Scale, respectively. Diet was assessed through Summary of Diabetes Self-Care Activities and Starting the Conversation tool. Incidence of hypoglycemic episodes was by patient report. Glycemic control was assessed with glycosylated hemoglobin (A1c) values from fingerstick blood sample. KEY RESULTS: The prevalence of food insecurity was 47.4 %. Chi-square tests revealed participants with food insecurity were more likely to be depressed (40.7 % vs. 15.4 %, p < 0.001), report diabetes distress (55.2 % vs. 33.8 %, p < 0.001) and have low medication adherence (52.9 % vs. 37.2 %, p = 0.02). Based on linear regression modeling, those with food insecurity had significantly higher mean A1c levels (ß = 0.51; p = 0.02) after adjusting for sex, age, race/ethnicity, language, education, marital status, BMI, insulin use, depression, diabetes distress and low medication adherence. CONCLUSIONS: Almost half of participants had food insecurity. Food insecurity was associated with depression, diabetes distress, low medication adherence and worse glycemic control. Even with adjustment, people with food insecurity had higher mean A1c levels than their food-secure counterparts, suggesting there may be other mediating factors, such as diet, that explain the relationship between food security status and diabetes control.

National patterns in diabetes screening: data from the National Health and Nutrition Examination Survey (NHANES) 2005-2012.

BACKGROUND: There are few current population-based estimates of the patterns of diabetes screening in the United States. The American Diabetes Association (ADA) recommends universal screening of adults ≥ 45 years, and high-risk adults < 45 years, but there is no current assessment of ADA guideline performance in detecting diabetes and prediabetes. Furthermore, data on racial/ethnic patterns of screening are limited. OBJECTIVE: Our aim was to estimate diabetes screening prevalence for the US adult population and specifically for those who meet ADA criteria; to report the prevalence of prediabetes and diabetes among these groups; and to determine if high-risk race/ethnicity was associated with reported screening. DESIGN: This was a Cross-sectional survey. PARTICIPANTS: Non-pregnant adults (≥ 21 years) without diabetes or prediabetes who participated in the National Health and Nutrition Examination Survey (NHANES) in 2005-2012 (n = 17,572) were included in the study. "Screening-recommended" participants, classified by ADA criteria, included (1) adults ≥ 45 years and (2) "high-risk" adults < 45 years. "Screening-not-recommended" participants were adults < 45 years who did not meet criteria. MAIN MEASURES: Diabetes screening status was obtained by self-report. We used calibrated HbA1c and/or fasting glucose levels to define undiagnosed diabetes and prediabetes. KEY RESULTS: Seventy-six percent of the study population (approximately 136 million US adults) met ADA criteria. Among them, less than half (46.2%) reported screening; undiagnosed diabetes affected 3.7% (5 million individuals), and undiagnosed prediabetes affected 36.3% (49 million people.) African Americans were more likely to report screening, both among adults ≥ 45 years and among "high risk" younger adults (OR 1.27 and 1.36, respectively.) Hispanic participants were also more likely to report screening (OR 1.31 for older adults, 1.42 for younger adults.) The screening rate among "screening-not-recommended" adults was 29.6%; the prevalence of diabetes and prediabetes were 0.4 and 10.2%, respectively. CONCLUSIONS: In a nationally representative sample, 76% of adults met ADA screening criteria, of whom fewer than half reported screening. Limitations include cross-sectional design and screening self-report.

An ABC transporter and an outer membrane lipoprotein participate in posttranslational activation of type VI secretion in Pseudomonas aeruginosa.

Pseudomonas aeruginosa is capable of injecting protein toxins into other bacterial cells through one of its three type VI secretion systems (T6SSs). The activity of this T6SS is tightly regulated on the posttranslational level by phosphorylation-dependent and -independent pathways. The phosphorylation-dependent pathway consists of a Threonine kinase/phosphatase pair (PpkA/PppA) that acts on a forkhead domain-containing protein, Fha1, and a periplasmic protein, TagR, that positively regulates PpkA. In the present work, we biochemically and functionally characterize three additional proteins of the phosphorylation-dependent regulatory cascade that controls T6S activation: TagT, TagS and TagQ. We show that similar to TagR, these proteins act upstream of the PpkA/PppA checkpoint and influence phosphorylation of Fha1 and, apparatus assembly and effector export. Localization studies demonstrate that TagQ is an outer membrane lipoprotein and TagR is associated with the outer membrane. Consistent with their homology to lipoprotein outer membrane localization (Lol) components, TagT and TagS form a stable inner membrane complex with ATPase activity. However, we find that outer membrane association of T6SS lipoproteins TagQ and TssJ1, and TagR, is unaltered in a ΔtagTS background. Notably, we found that TagQ is indispensible for anchoring of TagR to the outer membrane fraction. As T6S-dependent fitness of P. aeruginosa requires TagT, S, R and Q, we conclude that these proteins likely participate in a trans-membrane signalling pathway that promotes H1-T6SS activity under optimal environmental conditions.

Separate inputs modulate phosphorylation-dependent and -independent type VI secretion activation.

Productive intercellular delivery of cargo by secretory systems requires exquisite temporal and spatial choreography. Our laboratory has demonstrated that the haemolysin co-regulated secretion island I (HSI-I)-encoded type VI secretion system (H1-T6SS) of Pseudomonas aeruginosa transfers effector proteins to other bacterial cells. The activity of these effectors requires cell contact-dependent delivery by the secretion apparatus, and thus their export is highly repressed under planktonic growth conditions. Here we define regulatory pathways that orchestrate efficient secretion by this system. We identified a T6S-associated protein, TagF, as a posttranslational repressor of the H1-T6SS. Strains activated by TagF derepression or stimulated through a previously identified threonine phosphorylation pathway (TPP) share the property of secretory ATPase recruitment to the T6S apparatus, yet display different effector output levels and genetic requirements for their export. We also found that these two pathways respond to distinct stimuli; we identified surface growth as a physiological cue that activates the H1-T6SS exclusively through the TPP. Coordination of posttranslational triggering with cell contact-promoting growth conditions provides a mechanism for the T6SS to avoid wasteful release of effectors.

A minimalist substrate for enzymatic peptide and protein conjugation.

Recently a number of nonnatural prenyl groups containing alkynes and azides have been developed as handles to perform click chemistry on proteins and peptides ending in the sequence "CAAX", where C is a cysteine that becomes alkylated, A is an aliphatic amino acid and X is any amino acid. When such molecules are modified, a tag containing a prenyl analogue and the "CAAX box" sequence remains. Here we report the synthesis of an alkyne-containing substrate comprised of only nine nonhydrogen atoms. This substrate was synthesized in six steps from 3-methylbut-2-en-1-ol and has been enzymatically incorporated into both proteins and peptides by using protein farnesyltransferase. After prenylation the final three amino acids required for enzymatic recognition can be removed by using carboxypeptidase Y, leaving a single residue (the cysteine from the "CAAX box") and the prenyl analogue as the only modifications. We also demonstrate that this small tag minimizes the impact of the modification on the solubility of the targeted protein. Hence, this new approach should be useful for applications in which the presence of a large tag hinders the modified protein's solubility, reactivity, or utility.

Veteran peer Coaches Optimizing and Advancing Cardiac Health (Vet-COACH); design and rationale for a randomized controlled trial of peer support among Veterans with poorly controlled hypertension and other CVD risks.

BACKGROUND: Peer support can improve health for patients with chronic conditions; however, evidence for disease prevention is less clear and peer recruitment strategies are not well described. This paper describes a study protocol to evaluate a peer support intervention to improve hypertension control and reduce cardiovascular disease (CVD) risk. METHODS & RESEARCH DESIGN: Target enrollment for this two-site study is n = 400. Eligibility criteria include Veterans enrolled in Veterans Health Administration (VHA) primary care with poorly controlled hypertension and one other cardiovascular disease risk (smoking, overweight/obesity, or hyperlipidemia) who live in census tracts with high rates of hypertension. Enrolled participants are randomized to a home-based peer delivered self-management intervention (5 home visits and 5 phone calls with a peer health coach) versus usual care. The primary outcome is a change in systolic blood pressure (SBP) and secondary outcomes include change in CVD risk and health care use. RESULTS: Trial results are pending and participant enrollment is ongoing. We recruited peer coaches from Veterans who lived in census tracks with the highest rates of hypertension. To recruit Veteran peer coaches, we asked primary care providers (n = 41) and team nurses (n = 35) to nominate patients who they thought would be a good fit for the peer coach position (based on successful self-management and health care navigation) (n = 73 nominated from 964 patients). We interviewed 12 Veterans and trained 5 peer coaches. CONCLUSIONS: Results of this trial will inform peer support programs targeted to provide community-based delivery of prevention services to patients in high-risk areas. TRIAL REGISTRATION: Clinicaltrial.gov identifier NCT02697422 TRIAL STATUS: Enrollment for the randomized trial phase began in September 2017 and will be complete September 2019.

Methods Employed to Assess Weight Loss in Older Adults by Means of Electronic Medical Records: A Systematic Review.

Electronic medical records (EMRs) can be used to identify and categorize weight loss in older adults, but research has not scrutinized methods for doing so. Through a modified PRISMA protocol, we systematically reviewed published methods for quantifying weight change from EMRs. Articles (all available through July 2016) were identified through PubMed and SCOPUS searches, screened, and evaluated. We abstracted relevant data and tabulated the methods to assess weight change. The 13 selected articles showed little consistency in the approach to key methodological issues: 1) time ranges assessed; 2) removal of spurious values; 3) metrics to quantify weight change; 4) number of measures needed to estimate change; 5) threshold for significant weight change; and 6) relation to ideal weight. There was essentially no consensus around how to identify and categorize weight loss. Further investigation is needed to establish scientifically validated and clinically useful algorithms, accounting for the six issues above.

Associations between having an informal caregiver, social support, and self-care among low-income adults with poorly controlled diabetes.

Objective To determine whether the presence of an informal caregiver and the patient's level of social support are associated with better diabetes self-care among adults with poorly controlled diabetes. Methods Cross-sectional study using baseline data from 253 adults of age 30-70 with poorly controlled diabetes. Participants who reported receiving assistance with their diabetes from a friend or family member in the past month were classified as having a caregiver. We used multivariate linear and logistic regression models to evaluate the associations between having a caregiver and level of social support with five self-reported diabetes self-care behaviors: diet, foot checks, blood glucose monitoring, medications, and physical activity. Results Compared to participants with no informal caregiver, those with an informal caregiver were significantly more likely to report moderate or high medication adherence (OR = 1.93, 95% CI: 1.07-3.49, p = 0.028). When we included social support in the model, having a caregiver was no longer significantly associated with medication adherence (OR = 1.50, 95% CI: 0.80-2.82), but social support score was (OR = 1.22, 95% CI: 1.03-1.45, p = 0.023). Discussion Among low-income adults with poorly controlled diabetes, having both an informal caregiver and high social support for diabetes may have a beneficial effect on medication adherence, a key self-care target to improve diabetes control.

Peer Support for Achieving Independence in Diabetes (Peer-AID): design, methods and baseline characteristics of a randomized controlled trial of community health worker assisted diabetes self-management support.

BACKGROUND & OBJECTIVES: Community health workers (CHWs) may be an important mechanism to provide diabetes self-management to disadvantaged populations. We describe the design and baseline results of a trial evaluating a home-based CHW intervention. METHODS & RESEARCH DESIGN: Peer Support for Achieving Independence in Diabetes (Peer-AID) is a randomized, controlled trial evaluating a home-based CHW-delivered diabetes self-management intervention versus usual care. The study recruited participants from 3 health systems. Change in A1c measured at 12 months is the primary outcome. Changes in blood pressure, lipids, health care utilization, health-related quality of life, self-efficacy and diabetes self-management behaviors at 12 months are secondary outcomes. RESULTS: A total of 1438 patients were identified by a medical record review as potentially eligible, 445 patients were screened by telephone for eligibility and 287 were randomized. Groups were comparable at baseline on socio-demographic and clinical characteristics. All participants were low-income and were from diverse racial and ethnic backgrounds. The mean A1c was 8.9%, mean BMI was above the obese range, and non-adherence to diabetes medications was high. The cohort had high rates of co-morbid disease and low self-reported health status. Although one-third reported no health insurance, the mean number of visits to a physician in the past year was 5.7. Trial results are pending. CONCLUSIONS: Peer-AID recruited and enrolled a diverse group of low income participants with poorly controlled type 2 diabetes and delivered a home-based diabetes self-management program. If effective, replication of the Peer-AID intervention in community based settings could contribute to improved control of diabetes in vulnerable populations.

A type VI secretion system of Pseudomonas aeruginosa targets a toxin to bacteria.

The functional spectrum of a secretion system is defined by its substrates. Here we analyzed the secretomes of Pseudomonas aeruginosa mutants altered in regulation of the Hcp Secretion Island-I-encoded type VI secretion system (H1-T6SS). We identified three substrates of this system, proteins Tse1-3 (type six exported 1-3), which are coregulated with the secretory apparatus and secreted under tight posttranslational control. The Tse2 protein was found to be the toxin component of a toxin-immunity system and to arrest the growth of prokaryotic and eukaryotic cells when expressed intracellularly. In contrast, secreted Tse2 had no effect on eukaryotic cells; however, it provided a major growth advantage for P. aeruginosa strains, relative to those lacking immunity, in a manner dependent on cell contact and the H1-T6SS. This demonstration that the T6SS targets a toxin to bacteria helps reconcile the structural and evolutionary relationship between the T6SS and the bacteriophage tail and spike.