Improved functional abilities of the life-extended Drosophila mutant Methuselah are reversed at old age to below control levels

Methuselah (mth) is a chromosome 3 Drosophila mutant with an increased lifespan. A large number of studies have investigated the genetic, molecular, and biochemical mechanisms of the mth gene. Much less is known about the effects of mth on preservation of sensorimotor abilities throughout Drosophila’s lifespan, particularly in late life. The current study investigated functional senescence in mth and its parental-control line (w1118) in two experiments that measured age-dependent changes in flight functions and locomotor activity. In experiment 1, a total of 158 flies (81 mth and 77 controls) with an age range from 10 to 70 days were individually tethered under an infrared laser-sensor system that allowed monitoring of flight duration during phototaxic flight. We found that mth has a statistically significant advantage in maintaining continuous flight over control flies at age 10 days, but not during middle and late life. At age 70 days, the trend reversed and parental control flies had a small but significant advantage, suggesting an interaction between age and genotype in the ability to sustain flight. In experiment 2, a total of 173 different flies (97 mth and 76 controls) with an age range from 50 to 76 days were individually placed in a large well-lit arena (60 × 45 cm) and their locomotor activity quantified as the distance walked in a 1-min period. Results showed that mth flies had lower levels of locomotor activity relative to controls at ages 50 and 60 days. These levels converged for the two genotypes at the oldest ages tested. Findings show markedly different patterns of functional decline for the mth line relative to those previously reported for other life-extended genotypes, suggesting that different life-extending genes have dissimilar effects on preservation of sensory and motor abilities throughout an organism’s lifespan.     

Long-lived Indy induces reduced mitochondrial reactive oxygen species production and oxidative damage

Decreased Indy activity extends lifespan in D. melanogaster without significant reduction in fecundity, metabolic rate, or locomotion. To understand the underlying mechanisms leading to lifespan extension in this mutant strain, we compared the genome-wide gene expression changes in the head and thorax of adult Indy mutant with control flies over the course of their lifespan. A signature enrichment analysis of metabolic and signaling pathways revealed that expression levels of genes in the oxidative phosphorylation pathway are significantly lower in Indy starting at day 20. We confirmed experimentally that complexes I and III of the electron transport chain have lower enzyme activity in Indy long-lived flies by Day 20 and predicted that reactive oxygen species (ROS) production in mitochondria could be reduced. Consistently, we found that both ROS production and protein damage are reduced in Indy with respect to control. However, we did not detect significant differences in total ATP, a phenotype that could be explained by our finding of a higher mitochondrial density in Indy mutants. Thus, one potential mechanism by which Indy mutants extend life span could be through an alteration in mitochondrial physiology leading to an increased efficiency in the ATP/ROS ratio.     

Neuronal expression of Mgat1 rescues the shortened life span of Drosophila Mgat11 null mutants and increases life span

The enzyme UDP-GlcNAc:α3-D-mannoside β1,2-N-acetylglucosaminyltransferase I (GnT1, encoded by Mgat1) controls the synthesis of paucimannose N-glycans in Drosophila. We have previously reported that null mutations in Drosophila Mgat1 are viable but exhibit defects in locomotion, brain abnormalities, and a severely reduced life span. Here, we show that knockdown of Mgat1 in the central nervous system (CNS) of wild-type flies decreases locomotor activity and life span. This phenotype is similar to that observed in Drosophila Mgat1¹ null mutants, demonstrating that Mgat1 is required in the CNS. We also found that neuronal expression of a wild-type Mgat1 transgene rescued the shortened life span of Mgat1¹ null mutants and resulted in a dramatic 135% increase in mean life span relative to genetically identical controls. Neuronal expression of a wild-type Mgat1 transgene in wild-type flies resulted in a modest 9% increase in mean life span relative to genetically identical controls. In both Mgat1¹ null mutants and wild-type flies, neuronal expression of wild-type Mgat1 transgene resulted in a significant increase in GnT1 activity and resistance to oxidative stress. Whereas dietary restriction is not absolutely essential for the increased life span, it plays a role in the process. Interestingly, we observe a direct correlation between GnT1 activity and mean life span up to a maximum of ~136 days, showing that the ability of GnT1 activity to increase life span is limited. Altogether, these observations suggest that Mgat1-dependent N-glycosylation plays an important role in the control of Drosophila life span.     

Multiple-stress analysis for isolation of Drosophila longevity genes

Long-lived organisms tend to be more resistant to various forms of environmental stress. An example is the Drosophila longevity mutant, methuselah, which has enhanced resistance to heat, oxidants, and starvation. To identify genes regulated by these three stresses, we made a cDNA library for each by subtraction of "unstressed" from "stressed" cDNA and used DNA hybridization to identify genes that are regulated by all three. This screen indeed identified 13 genes, some already known to be involved in longevity, plus candidate genes. Two of these, hsp26 and hsp27, were chosen to test for their effects on lifespan by generating transgenic lines and by using the upstream activating sequence/GAL4 system. Overexpression of either hsp26 or hsp27 extended the mean lifespan by 30%, and the flies also displayed increased stress resistance. The results demonstrate that multiple-stress screening can be used to identify new longevity genes.     

The anti-aging effects of Ludwigia octovalvis on Drosophila melanogaster and SAMP8 mice

We investigated the anti-aging effects of Ludwigia octovalvis (Jacq.) P. H. Raven (Onagraceae), an extract of which is widely consumed as a healthful drink in a number of countries. Using the fruit fly, Drosophila melanogaster, as a model organism, we demonstrated that L. octovalvis extract (LOE) significantly extended fly lifespan on a high, but not a low, calorie diet, indicating that LOE may regulate lifespan through a dietary restriction (DR)-related pathway. LOE also attenuated age-related cognitive decline in both flies and in the senescence-accelerated-prone 8 (SAMP8) mouse, without causing any discernable negative trade-offs, including water intake, food intake, fecundity, or spontaneous motor activity. LOE contained high levels of polyphenols and flavonoids, which possess strong DPPH radical scavenging activity, and was shown to attenuate paraquat-induced oxidative damage and lethality in flies. Gas chromatography–mass spectrometry (GC-MS) analyses identified 17 known molecules, of which β-sitosterol and squalene were the two most abundant. We further demonstrated that β-sitosterol was capable of extending lifespan, likely through activating AMP-activated protein kinase (AMPK) in the fat body of adult flies. Taken together, our data suggest that LOE is a potent anti-aging intervention with potential for treating age-related disorders.

Electronic supplementary material

The online version of this article (doi:10.1007/s11357-013-9606-z) contains supplementary material, which is available to authorized users.     

Drosophila circadian rhythms in seminatural environments: Summer afternoon component is not an artifact and requires TrpA1 channels

Under standard laboratory conditions of rectangular light/dark cycles and constant warm temperature, Drosophila melanogaster show bursts of morning (M) and evening (E) locomotor activity and a “siesta” in the middle of the day. These M and E components have been critical for developing the neuronal dual oscillator model in which clock gene expression in key cells generates the circadian phenotype. However, under natural European summer conditions of cycling temperature and light intensity, an additional prominent afternoon (A) component that replaces the siesta is observed. This component has been described as an “artifact” of the TriKinetics locomotor monitoring system that is used by many circadian laboratories world wide. Using video recordings, we show that the A component is not an artifact, neither in the glass tubes used in TriKinetics monitors nor in open-field arenas. By studying various mutants in the visual and peripheral and internal thermo-sensitive pathways, we reveal that the M component is predominantly dependent on visual input, whereas the A component requires the internal thermo-sensitive channel transient receptor potential A1 (TrpA1). Knockdown of TrpA1 in different neuronal groups reveals that the reported expression of TrpA1 in clock neurons is unlikely to be involved in generating the summer locomotor profile, suggesting that other TrpA1 neurons are responsible for the A component. Studies of circadian rhythms under seminatural conditions therefore provide additional insights into the molecular basis of circadian entrainment that would otherwise be lost under the usual standard laboratory protocols.The circadian clock infiltrates almost every aspect of behavior and physiology of higher organisms and even some bacteria. Most studies of 24-h rhythms are carried out under strictly controlled laboratory conditions, an approach leading to a remarkably informative dissection of the clock, whose main molecular cogs are conserved among vertebrates and insects. Laboratory experiments are often extrapolated to the wild with the assumption that they reflect the natural situation. However, recent seminatural studies in mice, hamsters, and Drosophila have revealed some unexpected findings. For example, the widely held belief from laboratory studies that mice and golden hamsters are nocturnal needs to be revised because in the wild they are predominantly or exclusively diurnal (1, 2). Similarly in Drosophila melanogaster, locomotor rhythms studied in seminatural conditions reveal that deeply held, laboratory-derived assumptions may require significant revision. These include the crepuscular nature of fly activity, the role of the clock in “morning anticipation” and midday “siesta,” the requirement for clock gene expression in the central clock neurons for entrainment, and the role of light/dark (LD) cycles as the most important environmental Zeitgeber (“time giver”) in entraining the clock (3).Vanin et al. (3) observed that in the wild the phase of various features of circadian locomotor behavior such as the morning (M) and evening (E) components was best predicted by temperature, rather than “anticipation” of dawn and dusk over the seasonal LD cycle. In addition, at the warmer temperatures of European summers, flies did not generate an afternoon (A) siesta as in the laboratory. Instead, they dramatically increased their activity so that the major component of their locomotor profile was now the newly described A peak. The phase of the A component was modulated by mutation at the period (per) locus, suggesting that A represented a clock-mediated escape response from heat-induced stress (3, 4). Most surprisingly, null mutants of the negative regulators of the circadian clock period (per⁰¹) and timeless (tim⁰¹) exhibited naturally entrained behavioral profiles largely indistinguishable from those of wild-type strains. In sharp contrast, under laboratory conditions of constant 25 °C temperature and rectangular LD cycles, per⁰¹ and tim⁰¹ flies show no anticipation of dawn/dusk, and these mutants simply react to light-on or light-off signals with startle effects (5). The anticipatory nature of the M and E components in the laboratory led directly to the development of the dual oscillator model in the fly in which the Pigment Dispersing Factor (PDF) expressing s-LNv and l-LNvs (small and large lateral ventral neurons) generate the M locomotor component, whereas the dorsal lateral neurons, LNds, and dorsal neurons, DNs, produce the E component (6, 7).Although Vanin et al. focused predominantly on the phases of the major locomotor components under natural lighting and thermal conditions (3), in a similar natural study, Menegazzi et al. suggested that although per null mutants look similar in their behavioral phasing to wild type, the A peak tends to be larger in per⁰¹ mutants (4). These authors suggested that PER normally serves to reduce the amount of “inappropriate” activity that occurs during the warmest part of the day (4). Although their results were based on a very small sample of flies on a few days of recording, they were nevertheless welcomed in that they revealed that possessing a wild-type clock appeared to be behaviorally adaptive compared with having a severely disturbed clock.Another study performed under seminatural conditions at tropical latitudes has questioned the validity of the A component (8). These authors suggest instead that A represents a behavioral artifact as a result of flies avoiding the midday sun by sheltering in the shaded part of the glass activity tube where the TriKinetics infrared detectors are located, leading to inappropriate triggering of the sensor and high activity counts. In apparent support of this model, they observed that flies in open-field Petri dish arenas did not show an A component under summer conditions, although this interpretation has been criticized (9), in part because Petri dishes are well known to be problematic for Drosophila open-field behavioral recordings (10).Given the interest generated by Vanin et al. (3), we have revisited these natural studies and extended them with more sophisticated simulations of natural temperature and light cycles in the laboratory. By using video recordings of fly circadian activity in glass tubes and open-field arenas, we investigate whether the A component is an artifact. Furthermore, in both the Vanin et al. (3) and the Menegazzi et al. (4) studies, the classic per mutants were congenic with each other but were compared with three different wild-type strains so genetic background was not controlled. Using congenic controls we reexamine whether we can observe a phenotype for arrhythmic mutants in simulated seminatural conditions. Finally we study the A peak in a range of photoreceptor and thermoreceptor mutants to investigate the underlying genetic and neuroanatomical basis for this newly identified summer element of circadian behavior.     

Effect of melatonin in the antioxidant defense system in the locomotor muscles of the estuarine crab Neohelice granulata (Decapoda, Brachyura)

In vertebrates, many studies verified different effects of melatonin in the antioxidant defense system (ADS). In crustaceans, few studies have been conducted to verify this possibility. We verified the melatonin effects in the crab Neohelice granulata using low (0.002 and 0.02 pmol/crab) and high (2.0 and 20.0 pmol/crab) melatonin dosages in short-term (0.5 h) and long-term (9.5 h) experiments. We analyzed the antioxidant capacity against peroxyl radicals (ACAP), reactive oxygen species (ROS) concentration, levels of by products of lipid peroxidation (LPO), oxygen consumption (VO₂), the activity of glutamate cysteine ligase (γ-GCL) and catalase (CAT) and glutathione content (GSH). Finally, the effects of exogenous melatonin were verified in terms of melatonin and N¹-acetyl-N²-formyl-5-methoxykynuramine (AFMK) content in the muscles of N. granulata. In short-term experiment and low dosages, melatonin increased the VO₂, γ-GCL activity and GSH content (p < 0.05) and decreased melatonin content (p < 0.05) without effects in ROS, ACAP and LPO (p > 0.05). Possibly, melatonin is acting in the ADS increasing its efficiency and/or acting in mitochondrial activity and/or through signaling muscles to increase its consumption. AFMK was only detected in the eyestalk and cerebroid ganglia. In high dosages melatonin effects decreased, possibly by the desensitization of their receptors. In long-term experiment, melatonin decreased ACAP (p < 0.05), and CAT activity (p < 0.05) in low dosages. In high dosages melatonin reduced VO₂ (p < 0.05) and increased ACAP (p < 0.05), possibly stimulating others components of the ADS. In conclusion, melatonin in the locomotor muscles of N. granulata affects the antioxidant/pro-oxidant balance in a time and dosage dependent manner.     

Dyslipidemia-associated atherogenic oxidized lipids induce platelet hyperactivity through phospholipase Cγ2-dependent reactive oxygen species generation

Oxidized low-density lipoprotein (oxLDL) and associated oxidized phosphocholine-headgroup phospholipids (oxPCs) activate blood platelets through ligation of the scavenger receptor CD36. Previously, we found that oxLDL stimulated phosphorylation of phospholipase Cγ2 (PLCγ2). However, the functional relevance of PLCγ2 phosphorylation in oxLDL-mediated platelet hyperactivity remained elusive. Here, we set out to explore the functional importance of PLCγ2 in oxLDL-mediated platelet activation using human and genetically modified murine platelets. The CD36-specific oxidized phospholipid (oxPC_CD36) triggered the generation of reactive oxygen species (ROS) in platelets under static and arterial flow conditions. The ROS generation in response to oxPC_CD36 was sustained for up to 3 h but ablated in CD36- and PLCγ2-deficient platelets. The functional importance of ROS generation in response to atherogenic lipid stress was examined through measurement of P-selectin expression. OxPC_CD36 induced P-selectin expression, but required up to 60 min incubation, consistent with the timeline for ROS generation. P-selectin expression was not observed in CD36- and PLCγ2-deficient mice. The ability of oxPC_CD36 and oxLDL to stimulate P-selectin expression was prevented by incubation of platelets with the ROS scavenger N-acetyl-cysteine (NAC) and the NOX-2 inhibitor gp91ds-tat, but not with the NOX-1 inhibitor ML171. In summary, we provide evidence that prolonged exposure to oxLDL-associated oxidized phospholipids induces platelet activation via NOX-2-mediated ROS production in a CD36- and PLCγ2-dependent manner.     

The Parkinson's disease genes pink1 and parkin promote mitochondrial fission and/or inhibit fusion in Drosophila

Mutations in PTEN-induced kinase 1 (pink1) or parkin cause autosomal-recessive and some sporadic forms of Parkinson's disease. pink1 acts upstream of parkin in a common genetic pathway to regulate mitochondrial integrity in Drosophila. Mitochondrial morphology is maintained by a dynamic balance between the opposing actions of mitochondrial fusion, controlled by Mitofusin (mfn) and Optic atrophy 1 (opa1), and mitochondrial fission, controlled by drp1. Here, we explore interactions between pink1/parkin and the mitochondrial fusion/fission machinery. Muscle-specific knockdown of the fly homologue of Mfn (Marf) or opa1, or overexpression of drp1, results in significant mitochondrial fragmentation. Mfn-knockdown flies also display altered cristae morphology. Interestingly, knockdown of Mfn or opa1 or overexpression of drp1, rescues the phenotypes of muscle degeneration, cell death, and mitochondrial abnormalities in pink1 or parkin mutants. In the male germline, we also observe genetic interactions between pink1 and the testes-specific mfn homologue fuzzy onion, and between pink1 and drp1. Our data suggest that the pink1/parkin pathway promotes mitochondrial fission and/or inhibits fusion by negatively regulating mfn and opa1 function, and/or positively regulating drp1. However, pink1 and parkin mutant flies show distinct mitochondrial phenotypes from drp1 mutant flies, and flies carrying a heterozygous mutation in drp1 enhance the pink1-null phenotype, resulting in lethality. These results suggest that pink1 and parkin are likely not core components of the drp1-mediated mitochondrial fission machinery. Modification of fusion and fission may represent a novel therapeutic strategy for Parkinson's disease.     

Octopamine mediates starvation-induced hyperactivity in adult Drosophila

Starved animals often exhibit elevated locomotion, which has been speculated to partly resemble foraging behavior and facilitate food acquisition and energy intake. Despite its importance, the neural mechanism underlying this behavior remains unknown in any species. In this study we confirmed and extended previous findings that starvation induced locomotor activity in adult fruit flies Drosophila melanogaster. We also showed that starvation-induced hyperactivity was directed toward the localization and acquisition of food sources, because it could be suppressed upon the detection of food cues via both central nutrient-sensing and peripheral sweet-sensing mechanisms, via induction of food ingestion. We further found that octopamine, the insect counterpart of vertebrate norepinephrine, as well as the neurons expressing octopamine, were both necessary and sufficient for starvation-induced hyperactivity. Octopamine was not required for starvation-induced changes in feeding behaviors, suggesting independent regulations of energy intake behaviors upon starvation. Taken together, our results establish a quantitative behavioral paradigm to investigate the regulation of energy homeostasis by the CNS and identify a conserved neural substrate that links organismal metabolic state to a specific behavioral output.The CNS plays an essential role in energy homeostasis (1). It actively monitors changes in the internal energy state and modulates an array of physiological and behavioral responses to enable energy homeostasis. Foraging behavior is critical for the localization and acquisition of food supply and hence energy homeostasis. It has been extensively documented both in ethological settings (2, 3) and under well-controlled laboratory conditions (4). Laboratory rodents with limited food access exhibit stereotypic food anticipatory activity (FAA) several hours before the mealtime, which is characterized by a steady increase in locomotion and other appetitive behaviors (5). The neural substrate that drives FAA still remains elusive (5, 6). Notably, the regulation of FAA seems to be dissociable from that of feeding behavior (7, 8). These results hint at the presence of an independent and somewhat discrete regulatory mechanism of foraging behavior.Foraging behavior has also been extensively studied in invertebrate species such as the roundworm Caenorhabditis elegans (9) and fruit flies Drosophila melanogaster (10). Roundworm populations exhibit two naturally emerged foraging patterns: “solitary” worms disperse across the bacterial lawn, and “social” worms aggregate along the food edge and form clumps (9). This behavioral dimorphism is controlled by natural variations of the npr-1 (neuropeptide receptor resemblance) gene that encodes a receptor homologous to the receptor family of orexigenic neuropeptide Y in mammals (9). A comparable scenario has also been identified in larval fruit flies (10), with two distinct forms of foraging present in nature: “rover” and “sitter.” On food sources, sitter but not rover reduces moving speed for feeding (11). Natural variations of a single gene named foraging that encodes a cGMP-dependent protein kinase are responsible for this behavioral dimorphism (10). It remains unclear, however, whether the foraging strategies outlined above are driven by animals’ metabolic state, and if so whether npr-1 and foraging are involved (4). It is worth noting that both the roundworm and fruit fly larvae are continuous feeders. It is therefore difficult to disassociate the effect of the internal energy state on foraging behavior from that of acute change in food availability.In this present study, we sought to characterize foraging behavior in an intermittent feeder, the adult fruit fly. We confirmed that starved flies exhibited robust and sustained increase in their locomotor activity and provided evidence that it partly resembled foraging behavior. Furthermore, we found octopamine, a biological amine structurally related to vertebrate norepinephrine with similar physiological roles, both necessary and sufficient for starvation-induced hyperactivity. To summarize, our results reveal a highly conserved neural mechanism that promotes locomotion upon starvation, shedding important light on the regulation of foraging behavior by the CNS.     

Leishmania infantum DNA detection in Phlebotomus tobbi in a new northern focus of visceral leishmaniasis in Iran

Objective

To identify the vector(s), the parasite and the species composition of sand flies in the district during May-October 2012.

Methods

For reaching our objectives we used polymerase chain reaction of kDNA, ITS1-rDNA, followed by restriction fragment length polymorphism.

Results

Two species of Phlebotomus sergenti and Phlebotomus tobbi were the most prevalent among 8 species identified comprising 51.1% and 32.9% respectively. Among the 160 specimens of female sand flies tested by polymerase chain reaction of kDNA, ITS1-rDNA, followed by restriction fragment length polymorphisms, only 1 out of 80 Phlebotomus tobbi (1.25%) were positive to Leishmania infantum parasites.

Conclusions

Our finding showed that Phlebotomus tobbi may play as a vector to circulate the parasite of Leishmania infantum among reservoir(s) and human.     

Suppression of phase separation and giant enhancement of superconducting transition temperature in FeSe1?xTex thin films

We demonstrate the successful fabrication on CaF₂ substrates of FeSe_1−xTe_x films with 0 ≤ x ≤ 1, including the region of 0.1 ≤ x ≤ 0.4, which is well known to be the “phase-separation region,” via pulsed laser deposition that is a thermodynamically nonequilibrium method. In the resulting films, we observe a giant enhancement of the superconducting transition temperature, T_c, in the region of 0.1 ≤ x ≤ 0.4: The maximum value reaches 23 K, which is ∼1.5 times as large as the values reported for bulk samples of FeSe_1−xTe_x. We present a complete phase diagram of FeSe_1−xTe_x films. Surprisingly, a sudden suppression of T_c is observed at 0.1 < x < 0.2, whereas T_c increases with decreasing x for 0.2 ≤ x < 1. Namely, there is a clear difference between superconductivity realized in x = 0 ? 0.1 and in x ≥ 0.2. To obtain a film of FeSe_1−xTe_x with high T_c, the controls of the Te content x and the in-plane lattice strain are found to be key factors.Since the discovery of superconductivity in LaFeAs(O,F) (1), many studies concerning iron-based superconductors have been conducted. FeSe is the iron-based superconductor with the simplest crystal structure (2). The T_c of FeSe is ∼8 K, which is not very high in comparison with other iron-based superconductors. However, the value of T_c strongly depends on the applied pressure, and the temperature at which the resistivity becomes zero, Tczero, reaches as high as  ～ 30 K at 6 GPa (3). This suggests that FeSe samples with higher T_c are available by the fabrication of thin films because we can introduce lattice strain. Indeed, we have previously reported that FeSe films fabricated on CaF₂ substrates exhibit T_c values ∼1.5 times higher than those of bulk samples because of in-plane compressive strain (4). On the other hand, superconductivity with T_c of 65 K has recently been reported in a monolayer FeSe film on SrTiO₃ (5, 6). It is unclear whether this superconductivity results from the characteristics of the interface. However, this finding indicates that FeSe demonstrates potential as a very-high-T_c superconductor.The partial substitution of Te for Se in FeSe also raises T_c to a maximum of 14 K at x = 0.5 ? 0.6 (7). In FeSe_1−xTe_x, it is well known that we cannot obtain single-phase samples with 0.1 < x < 0.4 because of phase separation (7). Here, we focus on this region of phase separation. Generally, the process of film deposition involves crystal growth in a thermodynamically nonequilibrium state. Thus, film deposition provides an avenue for the synthesis of a material with a metastable phase. In this paper, we report the fabrication of epitaxial thin films of FeSe_1−xTe_x with 0 ≤ x ≤ 1 on CaF₂ substrates, using the pulsed laser deposition (PLD) method. We demonstrate that single-phase epitaxial films of FeSe_1−xTe_x with 0.1 ≤ x ≤ 0.4 are successfully obtained and that the maximum value of T_c is as large as 23 K, which is higher than the previously reported values for bulk and film samples of FeSe_1−xTe_x (7–12), except for those of the monolayer FeSe films (5, 6). Our results clearly show that the optimal Te content for the highest T_c for FeSe_1−xTe_x films on CaF₂ is different from the widely believed value for this system.Fig. 1A presents the X-ray diffraction patterns of FeSe_1−xTe_x films for x = 0 ? 0.5 on CaF₂. Here and hereafter, the Te content x of our films represents the nominal Te composition of the polycrystalline target. With the exception of an unidentified peak in the FeSe_0.5Te_0.5 film, only the 00l reflections of a tetragonal PbO-type structure are observed, which indicates that these films are well oriented along the c axis. Fig. 1 B and C presents enlarged segments of these plots near the 001 and 003 reflections, respectively. The 2θ values of the peak positions decrease with increasing x in a continuous manner, which is consistent with the fact that the c-axis length increases with increasing x. It should be noted that the values of the full widths at half maximum (FWHM), δ(2θ), of the FeSe_1−xTe_x films with x = 0.1 ? 0.4, which is known as the region of phase separation in the bulk samples (7), are δ(2θ) = 0.2°?0.3° for the 001 reflection and δ(2θ) = 0.4°?0.6° for the 003 reflection, which are nearly the same as the values for the FeSe and FeSe_0.5Te_0.5 films. This result is in sharp contrast to the previously reported result that the FWHM was broad only in films of FeSe_1−xTe_x with x = 0.1 and 0.3 (13), where phase separation has been believed to occur. The results presented in Fig. 1 A–C indicate the formation of a single phase in our FeSe_1−xTe_x films with x = 0.1 ? 0.4.Open in a separate windowFig. 1.(A) Out-of-plane X-ray diffraction patterns of FeSe_1−xTe_x thin films for x = 0 ? 0.5 with film thicknesses of 120–147 nm. The # symbols represent peaks associated with the substrate. The * represents an unidentified peak. Enlarged segments of the plots presented in A near the 001 and 003 peaks are shown in B and C, respectively. (D) The c-axis lengths of FeSe_1−xTe_x films, where the x value indicated on the horizontal axis is the nominal Te content. (E) Relations between the a-axis and c-axis lengths in FeSe_1−xTe_x films. The colors and shapes of the symbols correspond to the Te content x, as shown. The dashed lines are guides for the eye. The data for x = 0 and 0.5 presented in A–E are cited from refs. 4, 9, and 14.In Fig. 1D, the c-axis lengths of 29 films of FeSe_1−xTe_x are plotted as a function of x. The values of the c-axis lengths vary almost linearly with the nominal Te contents of the targets in the whole range of x, including both end-member materials. The evident formation of a single phase and the systematic change in the c-axis length strongly indicate that the nominal Te content of the polycrystalline target is nearly identical to that of the final FeSe_1−xTe_x film. Note that the compositional analysis of grown films using scanning electron microscopy/energy-dispersive X-ray (SEM/EDX) analysis is impossible for FeSe_1−xTe_x films on CaF₂ substrates because the energies of the K edge of Ca and the L edge of Te are very close to each other. The above-mentioned features indicate that phase separation is suppressed in our FeSe_1−xTe_x films with x = 0.1 ? 0.4 on CaF₂ substrates. To our knowledge, this result is the first manifestation of the suppression of phase separation in FeSe_1−xTe_x with x = 0.1 ? 0.4.Fig. 1E presents the relations between the a-axis and c-axis lengths in films of FeSe_1−xTe_x. At first glance, there seem to be no relations between the a-axis length and x, in sharp contrast to the behavior of the c-axis length. The a-axis and c-axis lengths of films with the same x show a weak negative correlation. This behavior cannot be explained by a difference in Te content of a film, which should result in a positive correlation. By contrast, if variations in c are caused by a difference in in-plane lattice strain, this behavior can be explained in terms of the Poisson effect. Indeed, the a-axis lengths of films of FeSe and FeSe_0.5Te_0.5 are smaller than those of bulk samples with the same composition. Thus, we consider that the a-axis length predominantly depends on the in-plane lattice strain rather than the Te content x. One might think that this behavior looks strange, because the lattice constant of CaF₂(aCaF2/2) is longer than the a of FeSe_1−xTe_x, which usually leads to a tensile strain. In a previous paper, the penetration of F⁻ ions from the CaF₂ substrates into the films was proposed as a possible mechanism for nontrivial compressive strain in FeSe_1−xTe_x films on CaF₂ substrates (15). Because of the smaller ionic radius of F⁻ than that of Se²⁻, this peculiar compressive strain can be explained by the partial substitution of F⁻ for Se²⁻ near the interface between a film and a substrate.Fig. 2 A–D presents the temperature dependences of the electrical resistivities, ρ, of 16 films of FeSe_1−xTe_x for x = 0.1 ? 0.4. The value of T_c depends on the film thickness, even in films with the same x. The highest Tconset, which is defined as the temperature where the electrical resistivity deviates from the normal-state behavior, and the Tczero of the FeSe_1−xTe_x films are 13.2 K and 11.5 K, respectively, for x = 0.1; 22.8 K and 20.5 K, respectively, for x = 0.2; 20.9 K and 19.9 K, respectively, for x = 0.3; and 20.9 K and 20.0 K, respectively, for x = 0.4. Compared with the results for bulk samples, a drastic enhancement of T_c is observed in these FeSe_1−xTe_x films. Surprisingly, the values of Tczero in the films with x = 0.2 and 0.4 exceed 20 K. These values are larger than those reported for FeSe_0.5Te_0.5 films (8, 10, 11, 14). In particular, the T_c of the FeSe_0.8Te_0.2 film with a thickness of 73 nm is ∼1.5 times as high as those of bulk crystals of FeSe_1−xTe_x with the optimal composition, x ≈ 0.5 (7). Based on the measurement of the ρ of the FeSe_0.8Te_0.2 film under a magnetic field applied along the c axis, we estimate an upper critical field at 0 K of μ₀H_c2 = 55.4 T, using the Werthamer–Helfand–Hohenberg (WHH) theory (16), which yields a Ginzburg–Landau coherence length at 0 K of ξ_ab(0) ～ 24.4 Å (SI Appendix). This value of μ₀H_c2 is approximately half the value for an FeSe_0.5Te_0.5 film on CaF₂ with a T_c of ∼16 K (9).Open in a separate windowFig. 2.Temperature dependences of the electrical resistivities, ρ, of FeSe_1−xTe_x thin films for (A) x = 0.1, (B) x = 0.2, (C) x = 0.3, and (D) x = 0.4 with different film thicknesses. Insets present enlarged views of the plots near the superconducting transition.Using the data shown above, we present the phase diagram of FeSe_1−xTe_x films on CaF₂ substrates in Fig. 3. For comparison, the data for bulk samples of FeSe_1−xTe_x (7, 17) are also plotted in Fig. 3. In bulk crystals, the optimal Te content to achieve the highest T_c is considered to be x ≈ 0.5, and phase separation occurs in the region of 0.1 ≤ x ≤ 0.4 (7). However, our data clearly demonstrate that this phase separation is absent and that the optimal composition for an FeSe_1−xTe_x film on a CaF₂ substrate is not x ≈ 0.5 but x ≈ 0.2. It should be noted that the dependence of T_c on x suddenly changes at the boundary defined by 0.1 < x < 0.2. Unlike the “dome-shaped” phase diagram that is familiar in iron-based superconductors, the values of T_c in films with 0.2 ≤ x ≤ 1 increase with decreasing x, whereas the strong suppression of T_c is observed at 0.1 < x < 0.2. The behavior in films with x ≥ 0.2 can be explained by the empirical law that shows the relation between T_c and structural parameters. In iron-based superconductors, it is well accepted that the bond angle of (Pn, Ch)-Fe-(Pn, Ch) (Pn, ?Pnictogen; Ch, ?Chalcogen), α (18, 19), and/or the anion height from the iron plane, h (20), are the critical structural parameters that determine the value of T_c. In bulk samples of FeSe_1−xTe_x, α and h approach their optimal values, i.e., α = 109.47° (18, 19) and h = 1.38 Å (20), with decreasing x (down to x = 0), which should be the same in FeSe_1−xTe_x films. Therefore, the increase of T_c in films with 0.2 ≤ x ≤ 1 with decreasing x can be explained by the optimization of α and/or h based on the empirical law. However, the sudden suppression of T_c in films with 0 ≤ x < 0.2 is not consistent with this scenario, and its origin should be sought among other factors. We consider there are two candidates for this origin from the structural analysis of bulk samples of FeSe_1−xTe_x. One is the effect of the orthorhombic distortion. In a bulk sample of FeSe, a structural phase transition from tetragonal to orthorhombic occurs at 90 K (21). However, in bulk samples of FeSe_1−xTe_x with x ～ 0.4 ? 0.6 where T_cs take optimum values, there are papers with different conclusions on the presence/absence of a similar type of structural transition to that of FeSe (22–24). It should be noted that a structural transition temperature is lower and that the orthorhombicity is much smaller than those of FeSe even in the report where the structural transition is present (24). These results on crystal structures suggest that the orthorhombic distortion results in a suppression of T_c. This scenario is applicable to the behavior of our films, if a large orthorhombic distortion is observed only in films with x = 0 ? 0.1. The other candidate is the change in the distance between the layers of Fe-Ch tetrahedra, δ. As shown in SI Appendix, in polycrystalline samples of FeSe_1−xTe_x, the δ value of FeSe is much smaller than those of FeSe_1−xTe_x with x ≥ 0.5 where δ is nearly independent of x (22). We speculate that the decrease of δ in FeSe is related with the suppression of T_c. Indeed, in polycrystalline samples, FeSe exhibits smaller values of δ and T_c than does FeSe_0.5Te_0.5 (7, 22), and the intercalation of alkali metals and alkaline earths into FeSe results in the c-axis length as large as ∼20 Å and T_c as high as 45 K (25, 26). At this moment, the origin of the suppression of T_c at 0.1 < x < 0.2 is unclear. Regardless of its origin, we believe that it is reasonable to distinguish between superconductivity in x = 0 ? 0.1 and in x ≥ 0.2. In other words, our phase diagram in Fig. 3 provides a previously unidentified view for superconductivity in FeSe_1−xTe_x, that is, a discontinuity in superconductivity of FeSe_1−xTe_x. We are able to come to this picture only after the data for x = 0.1 ? 0.4 become available in this study. If we remove a cause for the suppression of T_c in x=0,0.1 in some way, a further increase in T_c can be expected because of the optimization of structural parameters.Open in a separate windowFig. 3.Dependence of T_c on x. The red and blue circles represent the Tconset and Tczero values of the FeSe_1−xTe_x thin films, respectively. The black triangles represent the Tconset values obtained in measurements of the magnetic susceptibility of bulk samples (7, 17). The dashed curve is a guide for the eye.In conclusion, we prepared high-quality epitaxial thin films of FeSe_1−xTe_x on CaF₂ substrates, using the pulsed laser deposition method. We successfully obtained FeSe_1−xTe_x films with 0.1 ≤ x ≤ 0.4, which has long been considered to be the “phase-separation region,” using a thermodynamically nonequilibrium growth of film deposition. From the results of electrical resistivity measurements, a complete phase diagram is presented in this system, in which the maximum value of T_c is as high as 23 K at x = 0.2. Surprisingly, a sudden suppression of T_c is observed at 0.1 < x < 0.2, whereas T_c increases with decreasing x for 0.2 ≤ x < 1. This behavior is different from that of the dome-shaped phase diagram that is familiar in iron-based superconductors.     

Groups graded by root systems and property (T)

We establish property (T) for a large class of groups graded by root systems, including elementary Chevalley groups and Steinberg groups of rank at least 2 over finitely generated commutative rings with 1. We also construct a group with property (T) which surjects onto all finite simple groups of Lie type and rank at least two.Groups graded by root systems can be thought of as natural generalizations of Steinberg and Chevalley groups over rings. In recent preprints (1, 2), the authors of this paper determined a sufficient condition which almost implies property (T) for a group graded by a root system (see Theorem 1.1 below) and used this result to establish property (T) for Steinberg and Chevalley groups corresponding to reduced irreducible root systems of rank at least 2. The goal of this paper is to give an accessible exposition of those results and describe the main ideas used in their proofs.As will be shown in ref. 1, a substantial part of the general theory of groups graded by root systems can be developed using the term “root system” in a very broad sense. However, the majority of interesting examples (known to the authors) come from classical root systems (that is, finite crystallographic root systems), so in this paper we will only consider classical root systems (and refer to them simply as root systems).The basic idea behind the definition of a group graded by a root system Φ is that it should be generated by a family of subgroups indexed by Φ, which satisfies commutation relations similar to those between root subgroups of Chevalley and Steinberg groups.Definition: Let G be a group, Φ a root system, and {X_α}_α∈Φ a family of subgroups of G. We will say that the groups {X_α}_α∈Φ form a Φ-grading if for any α, β ∈ Φ with β ? ?α, the following inclusion holds:[X_α, X_β] ? ?X_γ:γ ∈ (?_≥1α + ?_≥1β)∩Φ?.[1.1]If in addition G is generated by the subgroups {X_α}, we will say that G is graded by Φ and that {X_α}_α∈Φ is a Φ-grading of G. The groups X_α themselves will be referred to as root subgroups.Clearly, the above definition is too general to yield any interesting structural results, and we are looking for the more restrictive notion of a strong grading. A sufficient condition for a Φ-grading to be strong is that the inclusion in [1.1] is an equality; however, requiring equality in general is too restrictive, as, for instance, it fails for Chevalley groups of type B_n over rings where 2 is not invertible. To formulate the definition of a strong grading in the general case, we need some additional terminology.Let Φ be a root system in a Euclidean space V with inner product ( ? ,  ? ). A subset B of Φ will be called Borel if B is the set of positive roots with respect to some system of simple roots Π of Φ. Equivalently, B is Borel if there exists v ∈ V, which is not orthogonal to any root in Φ, such that B = {γ ∈ Φ:(γ, v) > 0}.If B is a Borel subset and Π is the associated set of simple roots, the boundary of B, denoted by ??B, is the set of roots in B which are multiples of roots in Π. In particular, if Φ is reduced, we simply have ??B = Π.Definition: Let Φ be a root system, let G be a group and {X_α}_α∈Φ a Φ-grading of G. We will say that the grading {X_α} is strong if for any Borel subset B of Φ and any root γ ∈ B???B, we have X_γ ? ?X_β:β ∈ B?and?β ? ?γ?.It is easy to see that for any commutative ring R with 1 and any reduced irreducible root system Φ, the elementary Chevalley groups ??_Φ(R) and the Steinberg group St_Φ(R) are strongly graded by Φ.Our first main theorem asserts that any group strongly graded by an irreducible root system of rank at least 2 is in some sense close to having property (T).Theorem 1.1. (Ref. 1.) Let Φ be an irreducible root system of rank at least two, and let G be a group which admits a strong Φ-grading {X_α}. Then the union of root subgroups {X_α} is a Kazhdan subset of G (see section 1.2 for the definition).By definition, a group G has property (T) if it has a finite Kazhdan subset. Even though the root subgroups are almost never finite, Theorem 1.1 reduces proving property (T) for G to showing that the pair (G, ∪X_α) has relative property (T), and the latter can be achieved in many important examples. In particular, as a consequence of Theorem 1.1 and those results on relative property (T), we obtain the following theorem.Theorem 1.2. (Refs. 1 and 2.) Let Φ be a reduced irreducible root system of rank at least 2 and R a finitely generated ring (with 1). Assume that

• (a)R is commutative or
• (b)R is associative and Φ = A_n (with n ≥ 2) or
• (c)R is alternative and Φ = A₂.

Then the elementary Chevalley group ??_Φ(R) and the Steinberg group St_Φ(R) have property (T).Remark: Recall that a ring R is called alternative if (xx)y = x(xy) and x(yy) = (xy)y for all x, y ∈ R.Theorem 1.2b was previously established in ref. 3, but the result for root systems of types other than A was only known over rings of Krull dimension one (4). Later in the paper we will discuss several extensions of Theorem 1.2, dealing with twisted Steinberg groups over rings with involution.Convention: All rings in this paper are assumed to be unital and all groups are assumed to be discrete.     

Utilización de diferentes técnicas de biología molecular integradas en un algoritmo de identificación de micobacterias no tuberculosas

Introduction

The aim of the present work was to demonstrate the utility of a non-tuberculous mycobacteria (NTM) identification algorithm, which integrates different PCR-based techniques and basic phenotypic features. Moreover, the algorithm for pattern restriction analysis of hsp65 (hsp65 PRA) interpretation has been updated.

Methods

The workflow chosen consisted of the identification by a DNA hybridization probe method, followed by PCR-restriction enzyme analysis of hsp65 (hsp65 PRA) in those isolates that cannot be identified by hybridization probes. If necessary, 16S rRNA gene and hsp65 gene sequencing were used for speciation.

Results

A total of 236 NTM were collected, in which 102 (43.2%) isolates were identified by DNA specific probes and 76 (32.2%) isolates were identified with hsp65 PRA. Partial sequencing of the 16S rRNA gene was used for species identification of the remaining 58 (24.5%) isolates. Fifty-three (22.4%) were identified using this method. Five isolates (2.1%) were submitted for partial sequencing of hsp65 gene and one isolate was identified with this method. Four strains (1.7%) could not be identified at species level. Three new PRA patterns were found. Seven isolates tested positive with the AccuProbe Mycobacterium avium complex identification test but did not test positive with the M. avium or Mycobacterium intracellulare specific probes. Five and two of these isolates were identified as M. intracellulare and Mycobacterium colombiense, respectively.

Conclusion

This approach allowed us to identify almost all NTM isolates found in this study, including some recently described species.     

Oscillations in extracellular pH and reactive oxygen species modulate tip growth of Arabidopsis root hairs

Root hairs show highly localized cell expansion focused to their growing tips. This growth pattern is accomplished through restriction of secretion to the elongating apex and modulation of cell wall properties, with the wall just behind the tip becoming rigidified to resist the lateral expansive forces of turgor. In this report we show that root hairs exhibit oscillating growth that is associated with oscillating increases in extracellular pH and reactive oxygen species (ROS), which lag growth by ≈7 s. Consistent with a role for these changes in growth control, artificially increasing extracellular pH arrested root hair elongation, whereas decreasing pH elicited bursting at the tip. Similarly, application of exogenous ROS arrested elongation, whereas scavenging of ROS led to root hair bursting. Roots hairs of the root hair-defective rhd2-1 mutant, which lack a functional version of the NADPH oxidase ATRBOH C, burst at the transition to tip growth. This phenotype could be rescued by elevating the pH of the growth medium to ≥6.0. Such rescued root hairs showed reduced cytoplasmic ROS levels and a lack of the oscillatory production of ROS at the tip. However, they exhibited apparently normal tip growth, including generation of the tip-focused Ca²⁺ gradient thought to drive apical growth, indicating that ATRBOH C is not absolutely required to sustain tip growth. These observations indicate that root hair elongation is coupled to spatially distinct regulation of extracellular pH and ROS production that likely affect wall properties associated with the polarized expansion of the cell.