Adaptation of mycobacterium tuberculosis to biofilm growth is genetically linked to drug tolerance

Infections of Mycobacterium tuberculosis (MTB) require at least 6 months of multiple antibiotics for sterilization. This lengthy antibiotic regimen is widely attributed to a subpopulation of cells that acquire phenotypic tolerance to antibiotics. MTB readily forms pellicle biofilms at the air-media interface in vitro. These MTB biofilms contain more phenotypically antibiotic tolerant persister cells than cultures grown suspended in liquid medium (planktonic). Molecular mechanisms for the increase in persister frequency in MTB biofilms remain largely unknown. We utilized a high-throughput genomic approach (Tn-seq) to identify genes required by MTB to adapt to biofilm growth, but not planktonic growth, and to analyze their relationship with biofilm- associated stress and antibiotic tolerance. We identified multiple classes of mutants for formation of MTB pellicle biofilms. We hypothesized that the heterogeneous microenvironments of MTB biofilms create endogenous stressors that allow for self-selection of a population enriched for stress and antibiotic tolerant cells. Through use of a rifampicin (RIF)-hypersensitive mutant ΔpstC2-A1 strain that forms pellicle biofilms morphologically indistinguishable from wild-type (WT), we observed that intrinsic drug tolerance in constituent cells of biofilms determines the frequency of persisters: after 7 days of exposure to 50 µg/mL RIF, WT biofilms harbored approximately 20- fold more persisters than the mutant. These findings suggest that self-selection of tolerant cells during biofilm growth significantly promotes persister frequency. Using a transcriptomic analysisto characterize how constituent bacteria within MTB biofilms respond to environmental cues, we demonstrate biofilm-specific induction of the synthesis of isonitrile lipopeptides (INLP), which seem to be required for the development of MTB biofilm architecture based on mutant analysis. This work provides further insight into the antibiotic tolerant persistence of MTB biofilms, and identifies a biofilm-specific biomarker in INLPS for use in further investigation of this phenomenon. These findings provide molecular tools and potential antibiotic targets for investigation of MTB persisters, a significant public health obstacle to shortening the antibiotic treatment of TB

Stewards and Conservationists: Merging Moral Norms and the Theory of Planned Behavior to Understand Altruistic Conservation Behavior Among Hunters in Southwestern Utah

The leading cause of mortality in California Condors (condors) is lead poisoning, which occurs when condors ingest lead-based ammunition left in carcasses. As a critically endangered species with approximately 115 individuals remaining in the American southwest, increasing the adoption of non-lead ammunition is essential to the recovery of the species. In Utah, the Division of Wildlife Resources (DWR) uses communication with hunters as the primary tool for increasing the adoption of non-lead ammunition in southwestern Utah. In this research, we use social science theory and data collected from a survey of hunters throughout the region to develop a strategic communication framework aimed at increasing the use of non-lead ammunition among hunters. The strategic communication framework is intended to drive more specific, targeted, and effective messages regarding the use of non-lead ammunition by the Utah Division of Wildlife Resources and their conservation partners. Our findings suggest hunters, on average, hold positive attitudes towards non-lead ammunition and that their intention to use non-lead ammunition while hunting in in the area is high. This is good news for the DWR as an initial concern of the agency was that hunters may tend to hold negative attitudes towards non-lead ammunition and not be aware of the agency’s preferences and programs regarding the behavior. We asked hunters to self-report feelings of stewardship for the landscape and for the hunting tradition, believing that these feelings could be used as key leverage points to encourage the use of non-lead ammunition. These ‘personal norms’ were very strong amongst hunters; 92.7% of hunters consider themselves to be a steward of the natural landscape where they hunt and 88.4% believe they are stewards of the hunting tradition for future generations. Given these findings, we provide specific guidance on how the DWR can target personal norms in their communication with hunters regarding the use of non-lead ammunition. Tapping into feelings of stewardship over the landscape and family traditions are likely to be the most effective for effectuation behavioral change, reducing the use of lead ammunition, and conserving the condor population

Getting Lead Off the Landscape: A Theory and Data-Driven Approach to Increase Non-Lead Ammunition Use Among Hunters in the California Condor Range of Utah (USA)

The leading cause of mortality in California condors (condors) is lead poisoning, which occurs when condors ingest lead-based ammunition left in carcasses. As a critically endangered species with approximately 100 individuals remaining in the American southwest, increasing the adoption of non-lead ammunition is essential to the recovery of the species. In Utah, the Division of Wildlife Resources (DWR) uses communication with hunters as the primary tool for increasing the adoption of non-lead ammunition in southwestern Utah. In this research, we use social science theory and data collected from a survey of hunters throughout the region to develop a strategic communication framework aimed at increasing the use of non-lead ammunition among hunters. The strategic communication framework is intended to drive more specific, targeted, and effective messages regarding the use of non-lead ammunition by the Utah Division of Wildlife Resources and their conservation partners

Identifying Genetic Factors Promoting Fitness and Rifampicin Tolerance in Mycobacterium Tuberculosis Biofilms

OBJECTIVE: In 2011, the World Health Organization reported 8.7 million new cases of tuberculosis and 1.4 million tuberculosis-related deaths worldwide. The causative agent of this disease Mycobacterium tuberculosis is a notoriously persistent pathogen whose treatment requires a 6-9 month course of multiple antibiotics to clear the infection. Here, we used next generation DNA sequencing to identify several genetic factors involved in the tolerance of M. tuberculosis to the antibiotic rifampicin in the context of the biofilm, a common bacterial stress survival strategy. METHODS: A transposon insertion mutant library was constructed in the Erdman strain of M. tuberculosis. The library was then grown planktonically or in a biofilm and exposed to 5μg/ml of rifampicin. After exposure samples were processed and plated for colony formation. DNA was extracted from the colonies and prepared for sequencing by PCR amplification of transposon junction sites. DNA of these sites was sequenced using Illumina Hi-Seq technology and analyzed to find differential representation of transposon mutants between the unexposed and exposed library samples. RESULTS: Biofilms of M. tuberculosis are more tolerant than planktonic samples in vitro to rifampicin treatment. Seven genes at eight genomic positions were found to contain insertion sites that were ubiquitous in the samples unexposed to rifampicin, but found to be greatly decreased in the exposed biofilm samples: Rv0385, Rv1508c (at two positions), Rv1819c, Rv2779c, Rv3164c, Rv3796 and Rv3868. DISCUSSION: Further analysis of these mutants by using knockouts and studying the effects in vitro or in a mouse model will be necessary to confirm their role in rifampicin tolerance in an actual infection and determine any possible clinical benefits in exploiting these genetic factors. As latent infection remains a significant problem in global public health, exploitation of targets that contribute the persistence of infection could be a valuable tool in clearing infections

Henry Richards Correspondence

Entries include brief biographical information, a letter of correspondence from the Maine State Library, and a handwritten letter from his daughter

Incidental Malrotation in Adolescent Diagnosed with Appendicitis

17-year-old male patient with a history of autism presenting with right lower quadrant pain that started in the morning. The appendix was not able to be visualized by ultrasonography. An abdominal and pelvic CT with contrast showed a dilated appendix with surrounding inflammation. Malrotation was not identified on CT Scan. Patient was transferred to a nearby Children’s Hospital for laparoscopic appendectomy. During the procedure, the pediatric surgeon noted that initially, neither the appendix nor cecum was present in the right lower quadrant. This case represents an uncommon finding for a patient in the adolescent age group

Total cross sections

Regge theory provides a very simple and economical description of all total cross sectionsComment: 7 pages + 5 figures (appended as a ps file

Low SWAP Laminated Electrostatic Analyzers for CubeSat Applications

Instead of a cylindrical or spherical tophat design, these analyzers use flat plate with thousands of individual apertures. This flat plate geometry greatly eases manufacturing, and results in a very inexpensive instrument as describe by Enloe et al. (2003)

Crowdsourcing Crop Improvement in Sub?Saharan Africa: A Proposal for a Scalable and Inclusive Approach to Food Security

In sub?Saharan Africa, modern varieties released by the formal seed sector cover only a small part of the total crop acreage. Participatory approaches to crop improvement and seed production have been developed to overcome some of the barriers to modern variety development and seed distribution, but have not been widely scaled up. Crowdsourcing, such as seen in online citizen science projects, might inspire new approaches to upscale farmer?participatory seed innovation, specifically aiming at household food production. Using mobile technology, African farmers may be engaged in massively evaluating and distributing seeds. It is argued that a crowdsourcing approach to seed innovation would not only be scalable, but also inclusive through the strengthening of crop diversity as an open informational resource

Rosalind Richards Correspondence

Entries include brief biographical information, a typed biography, typed correspondence to Richards from the Maine State Library concerning her book, and a typed letter of presentation of a gift of books not for sale, written by her father