Tobacco Dependence Treatment Grants: A Collaborative Approach to the Implementation of WHO Tobacco Control Initiatives.

The number of global tobacco-related deaths is projected to increase from about 6 million to 8 million annually by 2030, with more than 80% of these occurring in low- and middle-income countries (LMICs). The World Health Organization Framework Convention on Tobacco Control (FCTC) came into force in 2005 and Article 14 relates specifically to the treatment of tobacco dependence. However, LMICs, in particular, face several barriers to implementing tobacco dependence treatment. This paper is a descriptive evaluation of a novel grant funding mechanism that was initiated in 2014 to address these barriers. Global Bridges. Healthcare Alliance for Tobacco Dependence Treatment aims to create and mobilize a global network of healthcare professionals and organizations dedicated to advancing evidence-based tobacco dependence treatment and advocating for effective tobacco control policy. A 2014 request for proposals (RFP) focused on these goals, particularly in LMICs, where funding for this work had been previously unavailable. 19 grants were awarded by Global Bridges to organizations in low- and middle-income countries across all six WHO regions. Virtually all focused on developing a tobacco dependence treatment curriculum for healthcare providers, while also influencing the political environment for Article 14 implementation. As a direct result of these projects, close to 9,000 healthcare providers have been trained in tobacco dependence treatment and an estimated 150,000 patients have been offered treatment. Because most of these projects are designed with a "train-the-trainer" component, two years of grant funding has been a tremendous catalyst for accelerating change in tobacco dependence treatment practices throughout the world. In order to foster such exponential growth and continue to maintain the impact of these projects, ongoing financial, educational, and professional commitments are required.

Varenicline for tobacco-dependence treatment in alcohol-dependent smokers: A randomized controlled trial.

BACKGROUND: Tobacco use is prevalent among persons with alcohol abuse and dependence. Varenicline has been shown to be the most effective pharmacotherapy for smoking cessation and may decrease alcohol consumption. The purpose of this study was to evaluate the efficacy of 12 weeks of varenicline for increasing smoking abstinence rates in smokers with alcohol abuse or dependence. METHODS: Participants were eligible for enrollment if they were 18 years or older, smoked 10 or more cigarettes per day for at least 6 months, had current alcohol abuse or dependence, and were interested in quitting smoking. Participants were randomly assigned to receive 12 weeks of varenicline 1 mg twice daily or matching placebo. The primary end point was 7-day point prevalence smoking abstinence at week 12. RESULTS: The 7-day point prevalence smoking abstinence rate at 12 weeks was significantly higher with varenicline (n = 16) than placebo (n = 17) (43.8% vs 5.9%; P = .01). At 24 weeks, the 7-day point prevalence smoking abstinence rate was still significantly higher with varenicline than placebo (31.3% vs 0%; P = .02). At 12 weeks, mean (SD) drinks per drinking day was significantly lower with varenicline than placebo (5.7 [3.9] vs 9.0 [5.3] drinks; treatment effect estimate, -2.8 [90% CI, -6.6 to -1.0]). Adverse events were minor and comparable to varenicline clinical trials. CONCLUSIONS: Varenicline is safe and efficacious for increasing smoking abstinence rates in smokers with alcohol abuse or dependence. Varenicline may decrease alcohol consumption in this population of smokers.

A Case Study into Microbial Genome Assembly Gap Sequences and Finishing Strategies.

This study characterized regions of DNA which remained unassembled by either PacBio and Illumina sequencing technologies for seven bacterial genomes. Two genomes were manually finished using bioinformatics and PCR/Sanger sequencing approaches and regions not assembled by automated software were analyzed. Gaps present within Illumina assemblies mostly correspond to repetitive DNA regions such as multiple rRNA operon sequences. PacBio gap sequences were evaluated for several properties such as GC content, read coverage, gap length, ability to form strong secondary structures, and corresponding annotations. Our hypothesis that strong secondary DNA structures blocked DNA polymerases and contributed to gap sequences was not accepted. PacBio assemblies had few limitations overall and gaps were explained as cumulative effect of lower than average sequence coverage and repetitive sequences at contig termini. An important aspect of the present study is the compilation of biological features that interfered with assembly and included active transposons, multiple plasmid sequences, phage DNA integration, and large sequence duplication. Our targeted genome finishing approach and systematic evaluation of the unassembled DNA will be useful for others looking to close, finish, and polish microbial genome sequences.

[The Mayo Clinic Nicotine Dependence Center]. / Centrum závislosti na nikotinu pri Mayo Clinic.

Since 1988, this world leading center provided treatment of tobacco dependence to tens of thousands of tobacco-dependent patients, educated thousands of health professionals and has rich research activities. Its system was a model for such similar centers including those in the Czech Republic.

Temporal Dynamics of In-Field Bioreactor Populations Reflect the Groundwater System and Respond Predictably to Perturbation.

Temporal variability complicates testing the influences of environmental variability on microbial community structure and thus function. An in-field bioreactor system was developed to assess oxic versus anoxic manipulations on in situ groundwater communities. Each sample was sequenced (16S SSU rRNA genes, average 10,000 reads), and biogeochemical parameters are monitored by quantifying 53 metals, 12 organic acids, 14 anions, and 3 sugars. Changes in dissolved oxygen (DO), pH, and other variables were similar across bioreactors. Sequencing revealed a complex community that fluctuated in-step with the groundwater community and responded to DO. This also directly influenced the pH, and so the biotic impacts of DO and pH shifts are correlated. A null model demonstrated that bioreactor communities were driven in part not only by experimental conditions but also by stochastic variability and did not accurately capture alterations in diversity during perturbations. We identified two groups of abundant OTUs important to this system; one was abundant in high DO and pH and contained heterotrophs and oxidizers of iron, nitrite, and ammonium, whereas the other was abundant in low DO with the capability to reduce nitrate. In-field bioreactors are a powerful tool for capturing natural microbial community responses to alterations in geochemical factors beyond the bulk phase.

Application of Long Sequence Reads To Improve Genomes for Clostridium thermocellum AD2, Clostridium thermocellum LQRI, and Pelosinus fermentans R7.

We and others have shown the utility of long sequence reads to improve genome assembly quality. In this study, we generated PacBio DNA sequence data to improve the assemblies of draft genomes for Clostridium thermocellum AD2, Clostridium thermocellum LQRI, and Pelosinus fermentans R7.

Development and Validation of Broad-Range Qualitative and Clade-Specific Quantitative Molecular Probes for Assessing Mercury Methylation in the Environment.

Two genes, hgcA and hgcB, are essential for microbial mercury (Hg) methylation. Detection and estimation of their abundance, in conjunction with Hg concentration, bioavailability, and biogeochemistry, are critical in determining potential hot spots of methylmercury (MeHg) generation in at-risk environments. We developed broad-range degenerate PCR primers spanning known hgcAB genes to determine the presence of both genes in diverse environments. These primers were tested against an extensive set of pure cultures with published genomes, including 13 Deltaproteobacteria, nine Firmicutes, and nine methanogenic Archaea genomes. A distinct PCR product at the expected size was confirmed for all hgcAB(+) strains tested via Sanger sequencing. Additionally, we developed clade-specific degenerate quantitative PCR (qPCR) primers that targeted hgcA for each of the three dominant Hg-methylating clades. The clade-specific qPCR primers amplified hgcA from 64%, 88%, and 86% of tested pure cultures of Deltaproteobacteria, Firmicutes, and Archaea, respectively, and were highly specific for each clade. Amplification efficiencies and detection limits were quantified for each organism. Primer sensitivity varied among species based on sequence conservation. Finally, to begin to evaluate the utility of our primer sets in nature, we tested hgcA and hgcAB recovery from pure cultures spiked into sand and soil. These novel quantitative molecular tools designed in this study will allow for more accurate identification and quantification of the individual Hg-methylating groups of microorganisms in the environment. The resulting data will be essential in developing accurate and robust predictive models of Hg methylation potential, ideally integrating the geochemistry of Hg methylation to the microbiology and genetics of hgcAB IMPORTANCE: The neurotoxin methylmercury (MeHg) poses a serious risk to human health. MeHg production in nature is associated with anaerobic microorganisms. The recent discovery of the Hg-methylating gene pair, hgcA and hgcB, has allowed us to design and optimize molecular probes against these genes within the genomic DNA for microorganisms known to methylate Hg. The protocols designed in this study allow for both qualitative and quantitative assessments of pure-culture or environmental samples. With these protocols in hand, we can begin to study the distribution of Hg-methylating organisms in nature via a cultivation-independent strategy.

Near-Complete Genome Sequence of the Cellulolytic Bacterium Bacteroides (Pseudobacteroides) cellulosolvens ATCC 35603.

We report the single-contig genome sequence of the anaerobic, mesophilic, cellulolytic bacterium, Bacteroides cellulosolvens. The bacterium produces a particularly elaborate cellulosome system, wherein the types of cohesin-dockerin interactions are opposite of other known cellulosome systems: cell-surface attachment is thus mediated via type-I interactions, whereas enzymes are integrated via type-II interactions.

Natural bacterial communities serve as quantitative geochemical biosensors.

UNLABELLED: Biological sensors can be engineered to measure a wide range of environmental conditions. Here we show that statistical analysis of DNA from natural microbial communities can be used to accurately identify environmental contaminants, including uranium and nitrate at a nuclear waste site. In addition to contamination, sequence data from the 16S rRNA gene alone can quantitatively predict a rich catalogue of 26 geochemical features collected from 93 wells with highly differing geochemistry characteristics. We extend this approach to identify sites contaminated with hydrocarbons from the Deepwater Horizon oil spill, finding that altered bacterial communities encode a memory of prior contamination, even after the contaminants themselves have been fully degraded. We show that the bacterial strains that are most useful for detecting oil and uranium are known to interact with these substrates, indicating that this statistical approach uncovers ecologically meaningful interactions consistent with previous experimental observations. Future efforts should focus on evaluating the geographical generalizability of these associations. Taken as a whole, these results indicate that ubiquitous, natural bacterial communities can be used as in situ environmental sensors that respond to and capture perturbations caused by human impacts. These in situ biosensors rely on environmental selection rather than directed engineering, and so this approach could be rapidly deployed and scaled as sequencing technology continues to become faster, simpler, and less expensive. IMPORTANCE: Here we show that DNA from natural bacterial communities can be used as a quantitative biosensor to accurately distinguish unpolluted sites from those contaminated with uranium, nitrate, or oil. These results indicate that bacterial communities can be used as environmental sensors that respond to and capture perturbations caused by human impacts.

Site-directed mutagenesis of HgcA and HgcB reveals amino acid residues important for mercury methylation.

Methylmercury is a potent neurotoxin that is produced by anaerobic microorganisms from inorganic mercury by a recently discovered pathway. A two-gene cluster, consisting of hgcA and hgcB, encodes two of the proteins essential for this activity. hgcA encodes a corrinoid protein with a strictly conserved cysteine proposed to be the ligand for cobalt in the corrinoid cofactor, whereas hgcB encodes a ferredoxin-like protein thought to be an electron donor to HgcA. Deletion of either gene eliminates mercury methylation by the methylator Desulfovibrio desulfuricans ND132. Here, site-directed mutants of HgcA and HgcB were constructed to determine amino acid residues essential for mercury methylation. Mutations of the strictly conserved residue Cys93 in HgcA, the proposed ligand for the corrinoid cobalt, to Ala or Thr completely abolished the methylation capacity, but a His substitution produced measurable methylmercury. Mutations of conserved amino acids near Cys93 had various impacts on the methylation capacity but showed that the structure of the putative "cap helix" region harboring Cys93 is crucial for methylation function. In the ferredoxin-like protein HgcB, only one of two conserved cysteines found at the C terminus was necessary for methylation, but either cysteine sufficed. An additional, strictly conserved cysteine, Cys73, was also determined to be essential for methylation. This study supports the previously predicted importance of Cys93 in HgcA for methylation of mercury and reveals additional residues in HgcA and HgcB that facilitate the production of this neurotoxin.

Impact of a countywide smoke-free workplace law on emergency department visits for respiratory diseases: a retrospective cohort study.

BACKGROUND: With the goal of reducing exposure to secondhand smoke, the state of Minnesota (MN), enacted a smoke-free law (i.e., Freedom to Breathe Act) in all workplaces, restaurants, and bars in 2007. This retrospective cohort study analyzes emergency department (ED) visits in Olmsted County, MN, for chronic obstructive pulmonary disease (COPD) and asthma over a five-year period to assess changes after enactment of the smoke-free law. METHODS: We calculated the rates of ED visits in Olmsted County, MN, with a primary diagnosis of COPD and asthma in the five-year period from January 1, 2005 to December 31, 2009. Analyses were performed using segmented Poisson regression to assess whether ED visit rates declined following enactment of the smoke free law after adjusting for potential underlying temporal trends in ED visit rates during this time period. RESULTS: Using segmented Poisson regression analyses, a significant reduction was detected in asthma-related ED visits (RR 0.814, p < 0.001) but not for COPD-related ED visits following the enactment of the smoke-free law. The reduction in asthma related ED visits was observed in both adults (RR 0.840, p = 0.015) and children (RR 0.751, p = 0.015). CONCLUSIONS: In Olmsted County, MN, asthma-related ED visits declined significantly after enactment of a smoke-free law. These results add to the body of literature supporting community health benefits of smoke-free policies in public environments and their potential to reduce health care costs.

Did we finally slay the evil dragon of cigarette smoking in the late 20th century?: unfortunately, the answer is no - the dragon is still alive and well in the 21st century and living in the third world. Shame on us!

If cigarettes were introduced as a new consumer product today, it is unlikely they would receive government regulatory approval. Cigarettes have proven biologic toxicities (carcinogenesis, atherogenesis, teratogenesis) and well-established causal links to human disease. Things were very different in 1913 when the R. J. Reynolds Tobacco Company introduced the first modern cigarette, the iconic Camel. By the early 1950s, definitive scientific reports linked cigarettes and human disease, but it was more than a half century later (2006) that cigarette manufacturers were found guilty by a federal court of deceptive product marketing regarding the health hazards of tobacco use. In the United States, cigarette smoking remains a major but slowly declining problem. But in developing countries, cigarette use is expanding tremendously. In global terms, the epidemic of smoking-caused disease is projected to increase rapidly in coming decades, not decline. Society may have begun to slowly win the smoking battle in the developed world, but we are resoundingly losing the global war on smoking. All is not lost! There is some good news! The 2003 Framework Convention on Tobacco Control, supported strongly by the American College of Chest Physicians, is the first global public health treaty of the new millennium. Many developed societies have begun planning to rid their countries of cigarettes in what is called the Endgame Strategy, and now is the time for the international medical community to help change tobacco policy to a worldwide endgame approach to rid all humanity of smoking-related diseases.

Improved yield of high molecular weight DNA coincides with increased microbial diversity access from iron oxide cemented sub-surface clay environments.

Despite over three decades of progress, extraction of high molecular weight (HMW) DNA from high clay soils or iron oxide cemented clay has remained challenging. HMW DNA is desirable for next generation sequencing as it yields the most comprehensive coverage. Several DNA extraction procedures were compared from samples that exhibit strong nucleic acid adsorption. pH manipulation or use of alternative ion solutions offered no improvement in nucleic acid recovery. Lysis by liquid N2 grinding in concentrated guanidine followed by concentrated sodium phosphate extraction supported HMW DNA recovery from clays high in iron oxides. DNA recovered using 1 M sodium phosphate buffer (PB) as a competitive desorptive wash was 15.22±2.33 µg DNA/g clay, with most DNA consisting of >20 Kb fragments, compared to 2.46±0.25 µg DNA/g clay with the Powerlyzer system (MoBio). Increasing PB concentration in the lysis reagent coincided with increasing DNA fragment length during initial extraction. Rarefaction plots of 16S rRNA (V1-V3 region) pyrosequencing from A-horizon and clay soils showed an ∼80% and ∼400% larger accessed diversity compared to the Powerlyzer soil DNA system, respectively. The observed diversity from the Firmicutes showed the strongest increase with >3-fold more operational taxonomic units (OTU) recovered.

Metallomics of two microorganisms relevant to heavy metal bioremediation reveal fundamental differences in metal assimilation and utilization.

Although as many as half of all proteins are thought to require a metal cofactor, the metalloproteomes of microorganisms remain relatively unexplored. Microorganisms from different environments are likely to vary greatly in the metals that they assimilate, not just among the metals with well-characterized roles but also those lacking any known function. Herein we investigated the metal utilization of two microorganisms that were isolated from very similar environments and are of interest because of potential roles in the immobilization of heavy metals, such as uranium and chromium. The metals assimilated and their concentrations in the cytoplasm of Desulfovibrio vulgaris strain Hildenborough (DvH) and Enterobacter cloacae strain Hanford (EcH) varied dramatically, with a larger number of metals present in Enterobacter. For example, a total of 9 and 19 metals were assimilated into their cytoplasmic fractions, respectively, and DvH did not assimilate significant amounts of zinc or copper whereas EcH assimilated both. However, bioinformatic analysis of their genome sequences revealed a comparable number of predicted metalloproteins, 813 in DvH and 953 in EcH. These allowed some rationalization of the types of metal assimilated in some cases (Fe, Cu, Mo, W, V) but not in others (Zn, Nd, Ce, Pr, Dy, Hf and Th). It was also shown that U binds an unknown soluble protein in EcH but this incorporation was the result of extracellular U binding to cytoplasmic components after cell lysis.

Combination varenicline and bupropion SR for tobacco-dependence treatment in cigarette smokers: a randomized trial.

IMPORTANCE: Combining pharmacotherapies for tobacco-dependence treatment may increase smoking abstinence. OBJECTIVE: To determine efficacy and safety of varenicline and bupropion sustained-release (SR; combination therapy) compared with varenicline (monotherapy) in cigarette smokers. DESIGN, SETTING, AND PARTICIPANTS: Randomized, blinded, placebo-controlled multicenter clinical trial with a 12-week treatment period and follow-up through week 52 conducted between October 2009 and April 2013 at 3 midwestern clinical research sites. Five hundred six adult (≥18 years) cigarette smokers were randomly assigned and 315 (62%) completed the study. INTERVENTIONS: Twelve weeks of varenicline and bupropion SR or varenicline and placebo. MAIN OUTCOMES AND MEASURES: Primary outcome was abstinence rates at week 12, defined as prolonged (no smoking from 2 weeks after the target quit date) abstinence and 7-day point-prevalence (no smoking past 7 days) abstinence. Secondary outcomes were prolonged and point-prevalence smoking abstinence rates at weeks 26 and 52. Outcomes were biochemically confirmed. RESULTS: At 12 weeks, 53.0% of the combination therapy group achieved prolonged smoking abstinence and 56.2% achieved 7-day point-prevalence smoking abstinence compared with 43.2% and 48.6% in varenicline monotherapy (odds ratio [OR], 1.49; 95% CI, 1.05-2.12; P = .03 and OR, 1.36; 95% CI, 0.95-1.93; P = .09, respectively). At 26 weeks, 36.6% of the combination therapy group achieved prolonged and 38.2% achieved 7-day point-prevalence smoking abstinence compared with 27.6% and 31.9% in varenicline monotherapy (OR, 1.52; 95% CI, 1.04-2.22; P = .03 and OR, 1.32; 95% CI, 0.91-1.91; P = .14, respectively). At 52 weeks, 30.9% of the combination therapy group achieved prolonged and 36.6% achieved 7-day point-prevalence smoking abstinence compared with 24.5% and 29.2% in varenicline monotherapy (OR, 1.39; 95% CI, 0.93-2.07; P = .11 and OR, 1.40; 95% CI, 0.96-2.05; P = .08, respectively). Participants receiving combination therapy reported more anxiety (7.2% vs 3.1%; P = .04) and depressive symptoms (3.6% vs 0.8%; P = .03). CONCLUSIONS AND RELEVANCE: Among cigarette smokers, combined use of varenicline and bupropion, compared with varenicline alone, increased prolonged abstinence but not 7-day point prevalence at 12 and 26 weeks. Neither outcome was significantly different at 52 weeks. Further research is required to determine the role of combination therapy in smoking cessation. TRIAL REGISTRATION: clinicaltrials.gov Identifier: http://clinicaltrials.gov/show/NCT00935818.

Mercury methylation by novel microorganisms from new environments.

Microbial mercury (Hg) methylation transforms a toxic trace metal into the highly bioaccumulated neurotoxin methylmercury (MeHg). The lack of a genetic marker for microbial MeHg production has prevented a clear understanding of Hg-methylating organism distribution in nature. Recently, a specific gene cluster (hgcAB) was linked to Hg methylation in two bacteria.1 Here we test if the presence of hgcAB orthologues is a reliable predictor of Hg methylation capability in microorganisms, a necessary confirmation for the development of molecular probes for Hg-methylation in nature. Although hgcAB orthologues are rare among all available microbial genomes, organisms are much more phylogenetically and environmentally diverse than previously thought. By directly measuring MeHg production in several bacterial and archaeal strains encoding hgcAB, we confirmed that possessing hgcAB predicts Hg methylation capability. For the first time, we demonstrated Hg methylation in a number of species other than sulfate- (SRB) and iron- (FeRB) reducing bacteria, including methanogens, and syntrophic, acetogenic, and fermentative Firmicutes. Several of these species occupy novel environmental niches for Hg methylation, including methanogenic habitats such as rice paddies, the animal gut, and extremes of pH and salinity. Identification of these organisms as Hg methylators now links methylation to discrete gene markers in microbial communities.

Draft genome sequences for three mercury-methylating, sulfate-reducing bacteria.

The genetic basis for bacterial mercury methylation has been described recently. For insights into the physiology of mercury-methylating bacteria, we present genome sequences for Desulfococcus multivorans strain DSM 2059, Desulfovibrio alkalitolerans strain DSM 16529, and Desulfovibrio species strain X2.

A randomized phase II clinical trial of high-dose nicotine patch therapy for smokeless tobacco users.

INTRODUCTION: Nicotine patch therapy has not been shown to be efficacious for increasing long-term (≥6 months) tobacco abstinence rates among smokeless tobacco (ST) users. Higher doses of nicotine patch therapy may be needed to increase tobacco abstinence rates in this population of tobacco users. METHODS: We randomized ST users who used ≥3 cans/pouches per week to either 8 weeks of high-dose nicotine patch therapy (42mg/day) or matching placebo patch. Subjects were followed for 6 months after randomization. RESULTS: Fifty-two subjects were randomized. Compared with placebo, high-dose nicotine patch therapy was associated with significantly higher prolonged tobacco abstinence at end-of-treatment (44% vs. 22%, odds ratio [OR] = 2.7, p = .050) and 3 months (40% vs. 19%, OR = 2.9, p = .047). High-dose nicotine patch therapy was associated with significant weight gain attenuation among tobacco abstinence subjects at 3 months (p = .013) and 6 months (p = .018). Compared with placebo, high-dose nicotine patch therapy was associated with nonsignificantly lower nicotine withdrawal scores. Adverse events were not significantly increased with high-dose nicotine patch therapy. CONCLUSIONS: High-dose nicotine patch therapy is safe and increases short-term tobacco abstinence rates among ST users who use ≥3 cans/pouches per week. High-dose nicotine patch therapy is associated with significant long-term attenuation of weight gain. Future studies to investigate the long-term efficacy of high-dose nicotine patch therapy and the comparative efficacy of this approach compared with standard nicotine patch doses for ST users seems warranted.