FNR-mediated regulation of bioluminescence and anaerobic respiration in the light-organ symbiont Vibrio fischeri

Vibrio fischeri induces both anaerobic respiration and bioluminescence during symbiotic infection. In many bacteria, the oxygen-sensitive regulator FNR activates anaerobic respiration, and a preliminary study using the light-generating lux genes from V. fischeri MJ1 cloned in Escherichia coli suggested that FNR stimulates bioluminescence. To test for FNR-mediated regulation of bioluminescence and anaerobic respiration in V. fischeri, we generated fnr mutants of V. fischeri strains MJ1 and ES114. In both strains, FNR was required for normal fumarate- and nitrate-dependent respiration. However, contrary to the report in transgenic E. coli, FNR mediated the repression of lux. ArcA represses bioluminescence, and ParcA-lacZ reporters showed reduced expression in fnr mutants, suggesting a possible indirect effect of FNR on bioluminescence via arcA. Finally, the fnr mutant of ES114 was not impaired in colonization of its host squid, Euprymna scolopes. This study extends the characterization of FNR to the Vibrionaceae and underscores the importance of studying lux regulation in its native background

A global map to aid the identification and screening of critical habitat for marine industries

Marine industries face a number of risks that necessitate careful analysis prior to making decisions on the siting of operations and facilities. An important emerging regulatory framework on environmental sustainability for business operations is the International Finance Corporation’s Performance Standard 6 (IFC PS6). Within PS6, identification of biodiversity significance is articulated through the concept of “Critical Habitat”, a definition developed by the IFC and detailed through criteria aligned with those that support internationally accepted biodiversity designations. No publicly available tools have been developed in either the marine or terrestrial realm to assess the likelihood of sites or operations being located within PS6-defined Critical Habitat. This paper presents a starting point towards filling this gap in the form of a preliminary global map that classifies more than 13 million km2 of marine and coastal areas of importance for biodiversity (protected areas, Key Biodiversity Areas [KBA], sea turtle nesting sites, cold- and warm-water corals, seamounts, seagrass beds, mangroves, saltmarshes, hydrothermal vents and cold seeps) based on their overlap with Critical Habitat criteria, as defined by IFC. In total, 5798×103 km2 (1.6%) of the analysis area (global ocean plus coastal land strip) were classed as Likely Critical Habitat, and 7526×103 km2 (2.1%) as Potential Critical Habitat; the remainder (96.3%) were Unclassified. The latter was primarily due to the paucity of biodiversity data in marine areas beyond national jurisdiction and/or in deep waters, and the comparatively fewer protected areas and KBAs in these regions. Globally, protected areas constituted 65.9% of the combined Likely and Potential Critical Habitat extent, and KBAs 29.3%, not accounting for the overlap between these two features. Relative Critical Habitat extent in Exclusive Economic Zones varied dramatically between countries. This work is likely to be of particular use for industries operating in the marine and coastal realms as an early screening aid prior to in situ Critical Habitat assessment; to financial institutions making investment decisions; and to those wishing to implement good practice policies relevant to biodiversity management. Supplementary material (available online) includes other global datasets considered, documentation and justification of biodiversity feature classification, detail of IFC PS6 criteria/scenarios, and coverage calculations

An expanded transposon mutant library reveals that Vibrio fischeri δ-aminolevulinate auxotrophs can colonize Euprymna scolopes

Libraries of defined mutants are valuable research tools but necessarily lack gene knockouts that are lethal under the conditions used in library construction. In this study, we augmented a Vibrio fischeri mutant library generated on a rich medium (LBS, which contains [per liter] 10 g of tryptone, 5 g of yeast extract, 20 g of NaCl, and 50 mM Tris [pH 7.5]) by selecting transposon insertion mutants on supplemented LBS and screening for those unable to grow on LBS. We isolated strains with insertions in alr, glr (murI), glmS, several heme biosynthesis genes, and ftsA, as well as a mutant disrupted 14 bp upstream of ftsQ. Mutants with insertions in ftsA or upstream of ftsQ were recovered by addition of Mg2+ to LBS, but their cell morphology and motility were affected. The ftsA mutant was more strongly affected and formed cells or chains of cells that appeared to wind back on themselves helically. Growth of mutants with insertions in glmS, alr, or glr was recovered with N-acetylglucosamine (NAG), D-alanine, or D-glutamate, respectively. We hypothesized that NAG, D-alanine, or D-glutamate might be available to V. fischeri in the Euprymna scolopes light organ; however, none of these mutants colonized the host effectively. In contrast, hemA and hemL mutants, which are auxotrophic for δ-aminolevulinate (ALA), colonized at wild-type levels, although mutants later in the heme biosynthetic pathway were severely impaired or unable to colonize. Our findings parallel observations that legume hosts provide Bradyrhizobium symbionts with ALA, but they contrast with virulence phenotypes of hemA mutants in some pathogens. The results further inform our understanding of the symbiotic light organ environment

Contribution of rapid evolution of the luxR-luxI intergenic region to the diverse bioluminescence outputs of Vibrio fischeri strains isolated from different environments

Vibrio fischeri serves as a valuable model of bacterial bioluminescence, its regulation, and its functional significance. Light output varies more than 10,000-fold in wild-type isolates from different environments, yet dim and bright strains have similar organization of the light-producing lux genes, with the activator-encoding luxR divergently transcribed from luxICDABEG. By comparing the genomes of bright strain MJ11 and the dimmer ES114, we found that the lux region has diverged more than most shared orthologs, including those flanking lux. Divergence was particularly high in the intergenic sequence between luxR and luxI. Analysis of the intergenic lux region from 18 V. fischeri strains revealed that, with one exception, sequence divergence essentially mirrored strain phylogeny but with relatively high substitution rates. The bases conserved among intergenic luxR-luxI sequences included binding sites for known regulators, such as LuxR and ArcA, and bases of unknown significance, including a striking palindromic repeat. By using this collection of diverse luxR-luxI regions, we found that expression of PluxI-lacZ but not PluxR-lacZ transcriptional reporters correlated with the luminescence output of the strains from which the promoters originated. We also found that exchange of a small stretch of the luxI-luxR intergenic region between two strains largely reversed their relative brightness. Our results show that the luxR-luxI intergenic region contributes significantly to the variable luminescence output among V. fischeri strains isolated from different environments, although other elements of strain backgrounds also contribute. Moreover, the lux system appears to have evolved relatively rapidly, suggesting unknown environment-specific selective pressures

Stimulant Medication and Reading Performance

The study examined the sustained effects of methylphenidate on reading performance in a sample of 42 boys, ages 8 to 11, with attention deficit-hyperactivity disorder (ADHD). Two subgroups were formed based on the presence or absence of co-occurring conduct disorders. Subjects were selected on the basis of their positive response to methylphenidate as determined in a series of original medication trials (Forness, Cantwell, Swanson, Hanna, & Youpa, 1991). For the purpose of this study, subjects were placed on their optimal dose of medication for a 6-week period and then tested on measures of oral reading and reading comprehension equivalent to those used in the original trials, retested after a week without medication (placebo), then tested again the following week after return to medication. Only the subgroup with conduct disorders responded, and this response was limited to reading comprehension improvement in only those subjects who also demonstrated improvement in oral reading on original trials. No response differences were found between subjects with or without learning disabilities.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/68569/2/10.1177_002221949202500205.pd

Rotationally driven magnetic reconnection in Saturn's dayside

Magnetic reconnection is a key process that explosively accelerates charged particles, generating phenomena such as nebular flares, solar flares and stunning aurorae. In planetary magnetospheres, magnetic reconnection has often been identified on the dayside magnetopause and in the nightside magnetodisc, where thin-current-sheet conditions are conducive to reconnection. The dayside magnetodisc is usually considered thicker than the nightside due to the compression of solar wind, and is therefore not an ideal environment for reconnection. In contrast, a recent statistical study of magnetic flux circulation strongly suggests that magnetic reconnection must occur throughout Saturn’s dayside magnetosphere. Additionally, the source of energetic plasma can be present in the noon sector of giant planetary magnetospheres. However, so far, dayside magnetic reconnection has only been identified at the magnetopause. Here, we report direct evidence of near-noon reconnection within Saturn’s magnetodisc using measurements from the Cassini spacecraft. The measured energetic electrons and ions (ranging from tens to hundreds of keV) and the estimated energy flux of ~2.6 mW m–2 within the reconnection region are sufficient to power aurorae. We suggest that dayside magnetodisc reconnection can explain bursty phenomena in the dayside magnetospheres of giant planets, which can potentially advance our understanding of quasi-periodic injections of relativistic electrons6 and auroral pulsations

Reconnection acceleration in Saturn's dayside magnetodisc:a multicase study with Cassini

Recently, rotationally driven magnetic reconnection was firstly discovered in Saturn’s dayside magnetosphere (Guo et al. 2018). This newly confirmed process could potentially drive bursty phenomena at Saturn, i.e., pulsating energetic particles and auroral emissions. Using Cassini’s measurements of magnetic fields and charged particles, we investigate particle acceleration features during three magnetic reconnection events observed in Saturn’s dayside magnetodisc. The results suggest that the rotationally driven reconnection process plays a key role in producing energetic electrons (up to 100 keV) and ions (several hundreds of keV). In particular, we find that energetic oxygen ions are locally accelerated at all three reconnection sites. Isolated, multiple reconnection sites were recorded in succession during an interval lasting for much less than one Saturn rotation period. Moreover, a secondary magnetic island is reported for the first time at the dayside, collectively suggesting that the reconnection process is not steady and could be ‘drizzle-like’. This study demonstrates the fundamental importance of internally driven magnetic reconnection in accelerating particles in Saturn’s dayside magnetosphere, and likewise in the rapidly rotating Jovian magnetosphere and beyond

Characterization of anticoagulant heparinoids by immunoprofiling

Heparinoids are used in the clinic as anticoagulants. A specific pentasaccharide in heparinoids activates antithrombin III, resulting in inactivation of factor Xa and–when additional saccharides are present–inactivation of factor IIa. Structural and functional analysis of the heterogeneous heparinoids generally requires advanced equipment, is time consuming, and needs (extensive) sample preparation. In this study, a novel and fast method for the characterization of heparinoids is introduced based on reactivity with nine unique anti-heparin antibodies. Eight heparinoids were biochemically analyzed by electrophoresis and their reactivity with domain-specific anti-heparin antibodies was established by ELISA. Each heparinoid displayed a distinct immunoprofile matching its structural characteristics. The immunoprofile could also be linked to biological characteristics, such as the anti-Xa/anti-IIa ratio, which was reflected by reactivity of the heparinoids with antibodies HS4C3 (indicative for 3-O-sulfates) and HS4E4 (indicative for domains allowing anti-factor IIa activity). In addition, the immunoprofile could be indicative for heparinoid-induced side-effects, such as heparin-induced thrombocytopenia, as illustrated by reactivity with antibody NS4F5, which defines a very high sulfated domain. In conclusion, immunoprofiling provides a novel, fast, and simple methodology for the characterization of heparinoids, and allows high-throughput screening of (new) heparinoids for defined structural and biological characteristics