The Retrohoming of Linear Group II Intron RNAs in Drosophila melanogaster Occurs by Both DNA Ligase 4–Dependent and –Independent Mechanisms

Mobile group II introns are bacterial retrotransposons that are thought to have invaded early eukaryotes and evolved into introns and retroelements in higher organisms. In bacteria, group II introns typically retrohome via full reverse splicing of an excised intron lariat RNA into a DNA site, where it is reverse transcribed by the intron-encoded protein. Recently, we showed that linear group II intron RNAs, which can result from hydrolytic splicing or debranching of lariat RNAs, can retrohome in eukaryotes by performing only the first step of reverse splicing, ligating their 3′ end to the downstream DNA exon. Reverse transcription then yields an intron cDNA, whose free end is linked to the upstream DNA exon by an error-prone process that yields junctions similar to those formed by non-homologous end joining (NHEJ). Here, by using Drosophila melanogaster NHEJ mutants, we show that linear intron RNA retrohoming occurs by major Lig4-dependent and minor Lig4-independent mechanisms, which appear to be related to classical and alternate NHEJ, respectively. The DNA repair polymerase θ plays a crucial role in both pathways. Surprisingly, however, mutations in Ku70, which functions in capping chromosome ends during NHEJ, have only moderate, possibly indirect effects, suggesting that both Lig4 and the alternate end-joining ligase act in some retrohoming events independently of Ku. Another potential Lig4-independent mechanism, reverse transcriptase template switching from the intron RNA to the upstream exon DNA, occurs in vitro, but gives junctions differing from the majority in vivo. Our results show that group II introns can utilize cellular NHEJ enzymes for retromobility in higher organisms, possibly exploiting mechanisms that contribute to retrotransposition and mitigate DNA damage by resident retrotransposons. Additionally, our results reveal novel activities of group II intron reverse transcriptases, with implications for retrohoming mechanisms and potential biotechnological applications

Selenium, Selenoenzymes, Oxidative Stress and Risk of Neoplastic Progression from Barrett's Esophagus: Results from Biomarkers and Genetic Variants

Clinical trials have suggested a protective effect of selenium supplementation on the risk of esophageal cancer, which may be mediated through the antioxidant activity of selenoenzymes. We investigated whether serum selenium concentrations, selenoenzyme activity, oxidative stress and genetic variation in selenoenzymes were associated with the risk of neoplastic progression to esophageal adenocarcinoma (EA) and two intermediate endpoints, aneuploidy and tetraploidy. In this prospective cohort study, during an average follow-up of 7.3 years, 47 EA cases, 41 aneuploidy cases and 51 tetraploidy cases accrued among 361 participants from the Seattle Barrett's Esophagus Research Study who were free of EA at the time of blood draw and had at least one follow-up visit. Development to EA was assessed histologically and aneuploidy and tetraploidy by DNA content flow cytometry. Serum selenium concentrations were measured using atomic absorption spectrometry, activity of glutathione peroxidase (GPX) 1 and GPX3 by substrate-specific coupled test procedures, selenoprotein P (SEPP1) concentrations and protein carbonyl content by ELISA method and malondialdehyde concentrations by HPLC. Genetic variants in GPX1-4 and SEPP1 were genotyped. Serum selenium was not associated with the risk of neoplastic progression to EA, aneuploidy or tetraploidy (P for trend = 0.25 to 0.85). SEPP1 concentrations were positively associated with the risk of EA [hazard ratio (HR) = 3.95, 95% confidence intervals (CI) = 1.42–10.97 comparing the third tertile with the first] and with aneuploidy (HR = 6.53, 95% CI = 1.31–32.58), but not selenoenzyme activity or oxidative stress markers. No genetic variants, overall, were associated with the risk of neoplastic progression to EA (global p = 0.12–0.69). Our results do not support a protective effect of selenium on risk of neoplastic progression to EA. Our study is the first to report positive associations of plasma SEPP1 concentrations with the risk of EA and aneuploidy, which warrants further investigation

Evidence for Limited Genetic Compartmentalization of HIV-1 between Lung and Blood

BACKGROUND:HIV-1 is frequently detected in the lungs of infected individuals and is likely important in the development of pulmonary opportunistic infections. The unique environment of the lung, rich in alveolar macrophages and with specialized local immune responses, may contribute to differential evolution or selection of HIV-1. METHODOLOGY AND FINDINGS:We characterized HIV-1 in the lung in relation to contemporaneous viral populations in the blood. The C2-V5 region of HIV-1 env was sequenced from paired lung (induced sputum or bronchoalveolar lavage) and blood (plasma RNA and proviral DNA from sorted or unsorted PBMC) from 18 subjects. Compartmentalization between tissue pairs was assessed using 5 established tree or distance-based methods, including permutation tests to determine statistical significance. We found statistical evidence of compartmentalization between lung and blood in 10/18 subjects, although lung and blood sequences were intermingled on phylogenetic trees in all subjects. The subject showing the greatest compartmentalization contained many nearly identical sequences in BAL sample, suggesting clonal expansion may contribute to reduced viral diversity in the lung in some cases. However, HIV-1 sequences in lung were not more homogeneous overall, nor were we able to find a lung-specific genotype associated with macrophage tropism in V3. In all four subjects in whom predicted X4 genotypes were found in blood, predicted X4 genotypes were also found in lung. CONCLUSIONS:Our results support a picture of continuous migration of HIV-1 between circulating blood and lung tissue, with perhaps a very limited degree of localized evolution or clonal replication

Targeted restoration of the intestinal microbiota with a simple, defined bacteriotherapy resolves relapsing Clostridium difficile disease in mice.

Relapsing C. difficile disease in humans is linked to a pathological imbalance within the intestinal microbiota, termed dysbiosis, which remains poorly understood. We show that mice infected with epidemic C. difficile (genotype 027/BI) develop highly contagious, chronic intestinal disease and persistent dysbiosis characterized by a distinct, simplified microbiota containing opportunistic pathogens and altered metabolite production. Chronic C. difficile 027/BI infection was refractory to vancomycin treatment leading to relapsing disease. In contrast, treatment of C. difficile 027/BI infected mice with feces from healthy mice rapidly restored a diverse, healthy microbiota and resolved C. difficile disease and contagiousness. We used this model to identify a simple mixture of six phylogenetically diverse intestinal bacteria, including novel species, which can re-establish a health-associated microbiota and clear C. difficile 027/BI infection from mice. Thus, targeting a dysbiotic microbiota with a defined mixture of phylogenetically diverse bacteria can trigger major shifts in the microbial community structure that displaces C. difficile and, as a result, resolves disease and contagiousness. Further, we demonstrate a rational approach to harness the therapeutic potential of health-associated microbial communities to treat C. difficile disease and potentially other forms of intestinal dysbiosis

Developing anti-tobacco messages for Australian Aboriginal and Torres Strait Islander peoples: evidence from a national cross-sectional survey

Background\ud Smoking rates in Australian Aboriginal and Torres Strait Islander peoples remain high, with limited impact of government measures for many subgroups. The aim of this cross-sectional study was to investigate differences in organisational practice for developing anti-tobacco messages for these target populations.\ud \ud Methods\ud Telephone interviews were conducted with 47 organisation representatives using a structured questionnaire based on health communication and health promotion frameworks. Responses were coded into phases of message development, message types (educational, threat, positive or advocacy), target groups, message recommendations, and evaluations undertaken. Cultural sensitivity for message development was divided into surface structure (use of images, language, demographics) and deep structure (use of socio-cultural values). A categorical principal component analysis explored the key dimensions of the findings and their component relationships.\ud \ud Results\ud Among organisations interviewed, a community-orientated, bottom-up approach for developing anti-tobacco messages was reported by 47% (n = 24); 55% based message development on a theoretical framework; 87% used a positive benefit appeal; 38% used threat messages. More Aboriginal Medical Services (AMSs) targeted youth (p < 0.005) and advised smokers to quit (p < 0.05) than other types of organisations. AMSs were significantly more likely to report using deep structure in tailoring messages compared with non-government (p < 0.05) and government organisations (p < 0.05). Organisations that were oriented to the general population were more likely to evaluate their programs (p < 0.05). A two-dimensional non-linear principal component analysis extracted components interpreted as "cultural understanding" (bottom-up, community-based approaches, deep structures) and "rigour" (theoretical frameworks, and planned/completed evaluations), and accounted for 53% of the variability in the data.\ud \ud Conclusion\ud Message features, associated with successful campaigns in other populations, are starting to be used for Aboriginal and Torres Strait Islander peoples. A model is proposed to facilitate the development of targeted anti-tobacco messages for Aboriginal and Torres Strait Islander peoples. Organisations could consider incorporating both components of cultural understanding-rigour to enable the growth of evidence-based practice

So Long and Thanks for All the Fish: Overexploitation of the Regionally Endemic Galapagos Grouper Mycteroperca olfax (Jenyns, 1840)

The regionally endemic Galapagos Grouper, locally known as bacalao, is one of the most highly prized finfish species within the Galapagos Marine Reserve (GMR). Concerns of overfishing, coupled with a lack of fishing regulations aimed at this species raises concerns about the current population health. We assessed changes in population health over a 30-year period using three simple indicators: (1) percentage of fish below reproductive size (L(m)); (2) percentage of fish within the optimum length interval (L(opt)); and (3) percentage of mega-spawners in the catch. Over the assessed period, none of the indicators reached values associated with healthy populations, with all indicators declining over time. Furthermore, the most recent landings data show that the vast majority of the bacalao caught (95.7%,) were below L(m), the number of fish within the L(opt) interval was extremely low (4.7%), and there were virtually no mega-spawners (0.2%). Bacalao fully recruit to the fishery 15 cm below the size at which 50% of the population matures. The Spawning Potential Ratio is currently 5% of potential unfished fecundity, strongly suggesting severe overfishing. Our results suggest the need for bacalao-specific management regulations that should include minimum (65 cm TL) and maximum (78 cm TL) landing sizes, slot limits (64–78 cm TL), as well as a closed season during spawning from October to January. It is recognized that these regulations are harsh and will certainly have negative impacts on the livelihoods of fishers in the short term, however, continued inaction will likely result in a collapse of this economically and culturally valuable species. Alternative sources of income should be developed in parallel with the establishment of fishing regulations to limit the socio-economic disruption to the fishing community during the transition to a more sustainable management regime