Importance of the Two Dissimilatory (Nar) Nitrate Reductases in the Growth and Nitrate Reduction of the Methylotrophic Marine Bacterium Methylophaga nitratireducenticrescens JAM1

Methylophaga nitratireducenticrescens JAM1 is the only reported Methylophaga species capable of growing under anaerobic conditions with nitrate as electron acceptor. Its genome encodes a truncated denitrification pathway, which includes two nitrate reductases, Nar1 and Nar2; two nitric oxide reductases, Nor1 and Nor2; and one nitrous oxide reductase, Nos; but no nitrite reductase (NirK or NirS). The transcriptome of strain JAM1 cultivated with nitrate and methanol under anaerobic conditions showed the genes for these enzymes were all expressed. We investigated the importance of Nar1 and Nar2 by knocking out narG1, narG2 or both genes. Measurement of the specific growth rate and the specific nitrate reduction rate of the knockout mutants JAM1ΔnarG1 (Nar1) and JAM1ΔnarG2 (Nar2) clearly demonstrated that both Nar systems contributed to the growth of strain JAM1 under anaerobic conditions, but at different levels. The JAM1ΔnarG1 mutant exhibited an important decrease in the nitrate reduction rate that consequently impaired its growth under anaerobic conditions. In JAM1ΔnarG2, the mutation induced a 20-h lag period before nitrate reduction occurred at specific rate similar to that of strain JAM1. The disruption of narG1 did not affect the expression of narG2. However, the expression of the Nar1 system was highly downregulated in the presence of oxygen with the JAM1ΔnarG2 mutant. These results indicated Nar1 is the major nitrate reductase in strain JAM1 but Nar2 appears to regulate the expression of Nar1

Technique de préparation des minéraux argileux en vue de l'analyse par diffraction des Rayons X et introduction à l'interprétation des diagrammes

Les techniques de préparation des échantillons pour l'analyse minéralogique des minéraux argileux sont présentées. Ces techniques consistent essentiellement à extraire une fraction fine (enrichie en argiles) à partir d'échantillons géologiques. Puis sont présentés les traitements de cette fraction enrichie en argiles en vue de leur analyse par diffraction des Rayons X. Enfin sont présentées les bases de la détermination des minéraux argileux par diffraction RX et l'interprétation des paramètres de cristallinité qui peuvent être extraits de ces diagrammes

Synergistic effects in mine offsite landscapes: Predicted ecosystem shifts could exacerbate mining effects on bryophyte community structure

Global change is shifting ecosystem type relative abundance in boreal forests, while the green energy transition results in increased mining activities around the globe. The interaction and consequent effects of these two trends on biodiversity have not been examined in depth. Bryophytes species can be used as indicators to measure these effects because they play key ecological roles in boreal forests. We identified and evaluated the interaction between ecosystem type (i.e., coniferous, deciduous, mixed forest and open canopy) and mining on microhabitat scale bryophyte diversity and composition in 1-km landscapes surrounding six mine sites at different stages of the mining lifecycle in the Canadian boreal forest. Irrespective of microhabitat type, the combined effects of ecosystem type and mining stage were interactive on bryophytes. Bryophyte richness and community compo-sition were negatively affected by offsite effects of mines in only deciduous and mixed forests. The interacted effects on bryophyte richness mainly occurred on the ground r microhabitats. We also found that deciduous, mixed forests (coniferous forest as a reference) and mines had a negative impact on the abundance of feather mosses and sphagna. Furthermore, indicator species were identified for areas affected by mines (Pohlia nutans and Dicranum polysetum) and for control areas (Sphagnum angustifolium and Plagiomnium cuspidatum). Our results suggest the predicted ecosystem shifts with global changes, from coniferous to deciduous forests, could poten-tially increase the effects of mining on forest ecosystem resistance through the changes in bryophyte community structure. Adding microhabitats (i.e., adding coarse woody debris) near mine sites is a potential strategy to maintain species richness. Collectively, these findings advance our understanding of how mining affects biodi-versity and highlight the importance of considering mine offsite landscapes in future environmental evaluations of development projects in the context of global changes

Étude d'une bactérie dénitrifiante du Biodôme de Montréal

Le nitrate est une molécule soluble qui diffuse rapidement dans les eaux de surfaces et les eaux souterraines, contaminant ainsi l’eau potable. Le nitrate contribue à l’eutrophisation des lacs et des rivières et devient toxique pour les animaux à partir de certaines concentrations. Methylophaga nitratireducticrescens JAM1 est une bactérie méthylotrophe isolée à partir d'un système de dénitrification d'eau de mer supplémenté en méthanol au Biodôme de Montréal. Cette bactérie compose à elle seule plus de 50% du biofilm dénitrifiant retrouvé dans le système de dénitrification. Contrairement aux autres espèces de Methylophaga, la souche JAM1 possède la particularité de croitre dans des conditions dénitrifiantes en présence de nitrate et de méthanol. La croissance de JAM1 dans les conditions dénitrifiantes, réduisant le nitrate en nitrite, est corrélée avec la présence de deux gènes codant pour des nitrate reductase narG. Le génome de JAM1 a été séquencé et consiste en un chromosome de 3,137,192 bp. Le séquençage du génome de JAM1 a confirmé la présence de deux opérons nar mais aussi de deux opérons nor (nitric oxide reductase) et d'un opéron nos (nitrous oxide reductase). De plus, une sequence nirK codant pour une nitrite reductase tronquée de 82 acides aminés a été trouvée, ce qui pourrait expliquer que la souche JAM1 est seulement capable de réduire le nitrate en nitrite. Notre travail consiste à étudier les gènes de dénitrification de la souche JAM1. Les deux gènes narG sont exprimés aussi bien en conditions aérobies que dénitrifiantes. Par ailleurs, les autres gènes liés à la dénitrification sont également exprimés, incluant le nirK tronqué. Des mesures d'expression ont montré que les gènes narG étaient exprimés différemment en fonction des conditions de cultures. Un mutant knockout de narG1 a montré de grosses différences de croissance et de taux de réduction du nitrate par rapport à la souche sauvage. Des différences d'expression du gène narG2 entre le mutant narG1 et la souche sauvage ont également été démontrées. La construction du mutant knockout narG2 est en cours et permettra de révéler l'importance de ce gène pour la croissance de la bactérie et la réduction du nitrate

A short-term assessment of nascent HIV-1 transmission clusters among newly diagnosed individuals using envelope sequence-based phylogenetic analyses

The identification of transmission clusters (TCs) of HIV-1 using phylogenetic analyses can provide insights into viral transmission network and help improve prevention strategies. We compared the use of partial HIV-1 envelope fragment of 1,070 bp with its loop 3 (108 bp) to determine its utility in inferring HIV-1 transmission clustering. Serum samples of recently (n = 106) and chronically (n = 156) HIV-1-infected patients with status confirmed were sequenced. HIV-1 envelope nucleotide-based phylogenetic analyses were used to infer HIV-1 TCs. Those were constructed using ClusterPickerGUI_1.2.3 considering a pairwise genetic distance of £10% threshold. Logistic regression analyses were used to examine the relationship between the demographic factors that were likely associated with HIV-1 clustering. Ninety-eight distinct consensus envelope sequences were subjected to phylogenetic analyses. Using a partial envelope fragment sequence, 42 sequences were grouped into 15 distinct small TCs while the V3 loop reproduces 10 clusters. The agreement between the partial envelope and the V3 loop fragments was significantly moderate with a Cohen’s kappa (j) coefficient of 0.59, p < .00001. The mean age (<38.8 years) and HIV-1 B subtype are two factors identified that were significantly associated with HIV-1 transmission clustering in the cohort, odds ratio (OR) = 0.25, 95% confidence interval (CI, 0.04–0.66), p = .002 and OR: 0.17, 95% CI (0.10–0.61), p = .011, respectively. The present study confirms that a partial fragment of the HIV-1 envelope sequence is a better predictor of transmission clustering. However, the loop 3 segment may be useful in screening purposes and may be more amenable to integration in surveillance programs

HIV-1 envelope sequence-based diversity measures for identifying recent infections

Identifying recent HIV-1 infections is crucial for monitoring HIV-1 incidence and optimizing public health prevention efforts. To identify recent HIV-1 infections, we evaluated and compared the performance of 4 sequence-based diversity measures including percent diversity, percent complexity, Shannon entropy and number of haplotypes targeting 13 genetic segments within the env gene of HIV-1. A total of 597 diagnostic samples obtained in 2013 and 2015 from recently and chronically HIV-1 infected individuals were selected. From the selected samples, 249 (134 from recent versus 115 from chronic infections) env coding regions, including V1-C5 of gp120 and the gp41 ectodomain of HIV-1, were successfully amplified and sequenced by next generation sequencing (NGS) using the Illumina MiSeq platform. The ability of the four sequence-based diversity measures to correctly identify recent HIV infections was evaluated using the frequency distribution curves, median and interquartile range and area under the curve (AUC) of the receiver operating characteristic (ROC). Comparing the median and interquartile range and evaluating the frequency distribution curves associated with the 4 sequence-based diversity measures, we observed that the percent diversity, number of haplotypes and Shannon entropy demonstrated significant potential to discriminate recent from chronic infections (p<0.0001). Using the AUC of ROC analysis, only the Shannon entropy measure within three HIV-1 env segments could accurately identify recent infections at a satisfactory level. The env segments were gp120 C2_1 (AUC = 0.806), gp120 C2_3 (AUC = 0.805) and gp120 V3 (AUC = 0.812). Our results clearly indicate that the Shannon entropy measure represents a useful tool for predicting HIV-1 infection recency

High-dose oral vitamin D supplementation and mortality in people aged 65-84 years: the VIDAL cluster feasibility RCT of open versus double-blind individual randomisation.

BACKGROUND: Randomised controlled trials demonstrating improved longevity are needed to justify high-dose vitamin D supplementation for older populations. OBJECTIVES: To demonstrate the feasibility of a large trial (n ≈ 20,000) of high-dose vitamin D in people aged 65-84 years through general practitioner (GP) practices, and to cluster randomise participating practices between open-label and double-blind randomisation to compare effects on recruitment, compliance and contamination. DESIGN: Twenty GP practices were randomised in matched pairs between open-label and double-blind allocation. Within each practice, patients were individually randomised to vitamin D or control (i.e. no treatment or placebo). Participants were invited to attend their GP practice to provide a blood sample and complete a lifestyle questionnaire at recruitment and again at 2 years. Randomisation by telephone followed receipt of a serum corrected calcium assay confirming eligibility ( 400 IU vitamin D per day at 2 years was 5.0% in open practices and 4.8% in double-blind practices. Mean serum 25(OH)D concentration was 51.5 nmol/l [95% confidence interval (CI) 50.2 to 52.8 nmol/l] with 82.6% of participants < 75 nmol/l at baseline. At 2 years, this increased to 109.6 nmol/l (95% CI 107.1 to 112.1 nmol/l) with 12.0% < 75 nmol/l in those allocated to vitamin D and was unaltered at 51.8 nmol/l (95% CI 49.8 to 53.8 nmol/l) in those allocated to no vitamin D (no treatment or placebo). CONCLUSIONS: A trial could recruit 20,000 participants aged 65-84 years through 200 GP practices over 2 years. Approximately 80% would be expected to adhere to allocated treatment (vitamin D or placebo) for 5 years. The trial could be conducted entirely by e-mail in participants aged < 80 years, but some participants aged 80-84 years would require postal follow-up. Recruitment and treatment compliance would be similar and contamination (self-administration of vitamin D) would be minimal, whether control participants are randomised openly to no treatment with no contact during the trial or randomised double-blind to placebo with monthly reminders. TRIAL REGISTRATION: Current Controlled Trials ISRCTN46328341 and EudraCT database 2011-003699-34. FUNDING: This project was funded by the National Institute for Health Research Health Technology Assessment programme and will be published in full in Health Technology Assessment; Vol. 24, No. 10. See the NIHR Journals Library website for further project information

HIV-1 envelope glycoprotein amino acids signatures associated with clade B transmitted/founder and recent viruses

Background: HIV-1 transmitted/founder viruses (TF) are selected during the acute phase of infection from a multitude of virions present during transmission. They possess the capacity to establish infection and viral dissemination in a new host. Deciphering the discrete genetic determinant of infectivity in their envelope may provide clues for vaccine design. Methods: One hundred twenty-six clade B HIV-1 consensus envelope sequences from untreated acute and early infected individuals were compared to 105 sequences obtained from chronically infected individuals using next generation sequencing and molecular analyses. Results: We identified an envelope amino acid signature associated with TF viruses. They are more likely to have an isoleucine (I) in position 841 instead of an arginine (R). This mutation of R to I (R841I) in the gp41 cytoplasmic tail (gp41CT), specifically in lentivirus lytic peptides segment 1 (LLP-1), is significantly enriched compared to chronic viruses (OR = 0.2, 95% CI (0.09, 0.44), p = 0.00001). Conversely, a mutation of lysine (K) to isoleucine (I) located in position six (K6I) of the envelope signal peptide was selected by chronic viruses and compared to TF (OR = 3.26, 95% CI (1.76–6.02), p = 0.0001). Conclusions: The highly conserved gp41 CT_ LLP-1 domain plays a major role in virus replication in mediating intracellular traffic and Env incorporation into virions in interacting with encoded matrix protein. The presence of an isoleucine in gp41 in the TF viruses’ envelope may sustain its role in the successful establishment of infection during the acute stage

Mineral magnetic characterization of the Upper Pleniglacial Nussloch loess sequence (Germany): an insight into local environmental processes

Presently, most loess/palaeosol magnetic susceptibility records are interpreted as following either the wind-vigour model or the pedogenic enhancement model. However redoxomorphic processes induced by waterlogging, often referred to gleying in the loess literature, are also known to alter loess deposits but their impact on loess/palaeosol magnetic susceptibility records has received little attention. The reported rock magnetic study aims to characterize the mineral magnetic response of loess to waterlogging-induced redoxomorphic processes, thus improving our understanding of mineral magnetic changes within loess deposits with respect to environmental and climate conditions. The Nussloch loess-palaeosol deposit (Rhine Valley, Germany) was targeted because it is one of the best-studied Pleniglacial deposits for Western Europe in which numerous tundra gley intervals have been identified. Moreover, a comprehensive high-resolution environmental magnetism study has never been undertaken for this site. Various rock magnetism experiments were conducted at both room and low temperatures to characterise the composition, concentration and relative magnetic grain size of the mineral magnetic assemblage. The relative changes in magnetic parameters within the investigated loess interval are primarily controlled by (1) varying concentrations of coarse-grained ferrimagnetic particles of detrital (aeolian) origin and (2) dissolution of fine-grained ferrimagnetic particles related to in situ post-depositional alteration promoted by waterlogging-induced redoxomorphic processes. Goethite is found to be ubiquitous throughout the studied interval and is argued to have both a primary (aeolian) and secondary (in situ) origin. We conclude, that redoxomorphic processes induced by waterlogging, if present, will hinder the interpretation of magnetic susceptibility variations within loess and palaeosol deposits following the expected relationships dictated by the wind-vigour and the pedogenic enhancement magnetism models