Elevated temperature differentially affects virulence, VirB protein accumulation, and T-pilus formation in different Agrobacterium tumefaciens and Agrobacterium vitis strains.

That gene transfer to plant cells is a temperature-sensitive process has been known for more than 50 years. Previous work indicated that this sensitivity results from the inability to assemble a functional T pilus required for T-DNA and protein transfer to recipient cells. The studies reported here extend these observations and more clearly define the molecular basis of this assembly and transfer defect. T-pilus assembly and virulence protein accumulation were monitored in Agrobacterium tumefaciens strain C58 at different temperatures ranging from 20 degrees C to growth-inhibitory 37 degrees C. Incubation at 28 degrees C but not at 26 degrees C strongly inhibited extracellular assembly of the major T-pilus component VirB2 as well as of pilus-associated protein VirB5, and the highest amounts of T pili were detected at 20 degrees C. Analysis of temperature effects on the cell-bound virulence machinery revealed three classes of virulence proteins. Whereas class I proteins (VirB2, VirB7, VirB9, and VirB10) were readily detected at 28 degrees C, class II proteins (VirB1, VirB4, VirB5, VirB6, VirB8, VirB11, VirD2, and VirE2) were only detected after cell growth below 26 degrees C. Significant levels of class III proteins (VirB3 and VirD4) were only detected at 20 degrees C and not at higher temperatures. Shift of virulence-induced agrobacteria from 20 to 28 or 37 degrees C had no immediate effect on cell-bound T pili or on stability of most virulence proteins. However, the temperature shift caused a rapid decrease in the amount of cell-bound VirB3 and VirD4, and VirB4 and VirB11 levels decreased next. To assess whether destabilization of virulence proteins constitutes a general phenomenon, levels of virulence proteins and of extracellular T pili were monitored in different A. tumefaciens and Agrobacterium vitis strains grown at 20 and 28 degrees C. Levels of many virulence proteins were strongly reduced at 28 degrees C compared to 20 degrees C, and T-pilus assembly did not occur in all strains except "temperature-resistant" Ach5 and Chry5. Virulence protein levels correlated well with bacterial virulence at elevated temperature, suggesting that degradation of a limited set of virulence proteins accounts for the temperature sensitivity of gene transfer to plants.

Temperature and precipitation effects on δ13C depth profiles in SOM under temperate beech forests

Enrichment of 13C in SOM with soil depth is related to interacting processes influenced by temperature and precipitation. Our objectives were to derive climate effects on patterns of vertical δ13C values of soil organic matter (SOM) while minimizing the effect of confounding variables. We investigated vertical changes in δ13C values of SOM in 1-cm depth intervals in silvicultural mature beech (Fagus sylvatica L.) forest ecosystems in northern Rhineland-Palatinate across gradients of MAT (7.9 to 9.7 °C mean annual temperature) and MAP (607 to 1085 mm mean annual precipitation) in winter 2011. Forest stands (n = 10) were chosen based on data sets provided by the Rhineland-Palatinate Forest Administration so that variations in these gradients occurred while other environmental factors like physico-chemical soil properties, tree species, stand age, exposition and precipitation (for the temperature gradient) or temperature (for the precipitation gradient) did not differ among study sites. From litter down to the mineral soil at 10 cm depth, soil organic carbon (SOC) content decreased (47.5 ± SE 0.1% to 2.5 ± 0.1%) while the δ13C values increased (− 29.4 ± 0.1‰ to − 26.1 ± 0.1‰). Litter of sites under higher MAP/lower MAT had lower δ13C values which was in line with literature data on climate driven plant physiological process. To compare the dimension of the vertical 13C enrichment, δ13C values were regressed linearly against log-transformed carbon contents yielding absolute values of these slopes (beta). Beta values ranged between 0.6 and 4.5 (range of r from − 0.7 to − 1.0; p < 0.01). Due to an assumed decay continuum and similar variations of δ13C values in litter and in 10 cm depth, we conclude that effects on isotope composition in the Oi layer continue vertically and therefore, δ13C values in litter do not solely control beta values. Beta values decreased with increasing MAT (r = − 0.83; p < 0.05). Reduced soil moisture and therefore both, reduced microbial activity and reduced downward transport of microbial cycled DOM (=13C enriched) might be responsible for less pronounced δ13C depth profiles in case of high temperatures. Greater C:N ratios (lower degradability) of the litter under higher temperatures likely contributed to these depth trends. Beta values increased with increasing MAP (r = 0.73; p < 0.05). We found decreasing C:N ratios in the mineral soil that possibly indicates higher decomposition under higher precipitation. Exclusion of the organic layers from linear regressions indicated a stronger impact of MAP on the development of δ13C depth profiles. Our results confirm temperature and precipitation effects on δ13C depth profiles and indicate stronger 13C enrichment under lower MAT/higher MAP. Therefore, time series of vertical δ13C depth profiles might provide insights into climate change effects.

Electrical resistivity change in amorphous Ta42Si13N45 films by stress relaxation

In a first experiment, a reactively sputtered amorphous Ta₄₂Si₁₃N₄₅ film about 260 nm thick deposited on a flat and smooth alumina substrate was thermally annealed in air for 30 min and let cooled again repeatedly at successively higher temperatures from 200 to 500 °C. This treatment successively and irreversibly increases the room temperature resistivity of the film monotonically from its initial value of 670 μΩ cm to a maximum of 705 μΩ cm (+5.2 %). Subsequent heat treatments at temperatures below 500 °C and up to 6 h have no further effect on the room temperature resistivity. The new value remains unchanged after 3.8 years of storage at room temperature. In a second experiment, the evolution of the initially compressive stress of a film similarly deposited by reactive sputtering on a 2-inch silicon wafer was measured by tracking the wafer curvature during similar thermal annealing cycles. A similar pattern of irreversible and reversible changes of stress was observed as for the film resistivity. Transmission electron micrographs and secondary ion mass profiles of the film taken before and after thermal annealing in air establish that both the structure and the composition of the film scarcely change during the annealing cycles. We reason that the film stress is implicated in the resistivity change. In particular, to interpret the observations, a model is proposed where the interface between the film and the substrate is mechanically unyielding.

Murine coronavirus ubiquitin-like domain is important for papain-like protease stability and viral pathogenesis

Ubiquitin-like domains (Ubls) now are recognized as common elements adjacent to viral and cellular proteases; however, their function is unclear. Structural studies of the papain-like protease (PLP) domains of coronaviruses (CoVs) revealed an adjacent Ubl domain in severe acute respiratory syndrome CoV, Middle East respiratory syndrome CoV, and the murine CoV, mouse hepatitis virus (MHV). Here, we tested the effect of altering the Ubl adjacent to PLP2 of MHV on enzyme activity, viral replication, and pathogenesis. Using deletion and substitution approaches, we identified sites within the Ubl domain, residues 785 to 787 of nonstructural protein 3, which negatively affect protease activity, and valine residues 785 and 787, which negatively affect deubiquitinating activity. Using reverse genetics, we engineered Ubl mutant viruses and found that AM2 (V787S) and AM3 (V785S) viruses replicate efficiently at 37°C but generate smaller plaques than wild-type (WT) virus, and AM2 is defective for replication at higher temperatures. To evaluate the effect of the mutation on protease activity, we purified WT and Ubl mutant PLP2 and found that the proteases exhibit similar specific activities at 25°C. However, the thermal stability of the Ubl mutant PLP2 was significantly reduced at 30°C, thereby reducing the total enzymatic activity. To determine if the destabilizing mutation affects viral pathogenesis, we infected C57BL/6 mice with WT or AM2 virus and found that the mutant virus is highly attenuated, yet it replicates sufficiently to elicit protective immunity. These studies revealed that modulating the Ubl domain adjacent to the PLP reduces protease stability and viral pathogenesis, revealing a novel approach to coronavirus attenuation. IMPORTANCE Introducing mutations into a protein or virus can have either direct or indirect effects on function. We asked if changes in the Ubl domain, a conserved domain adjacent to the coronavirus papain-like protease, altered the viral protease activity or affected viral replication or pathogenesis. Our studies using purified wild-type and Ubl mutant proteases revealed that mutations in the viral Ubl domain destabilize and inactivate the adjacent viral protease. Furthermore, we show that a CoV encoding the mutant Ubl domain is unable to replicate at high temperature or cause lethal disease in mice. Our results identify the coronavirus Ubl domain as a novel modulator of viral protease stability and reveal manipulating the Ubl domain as a new approach for attenuating coronavirus replication and pathogenesis.

Late Pliocene and Early Pleistocene dinoflagellate cysts and sea surface temperture reconstruction for several IODP and DSDP sites in the North Atlantic

In an attempt to document the palaeoecological affinities of individual extant and extinct dinoflagellate cysts, Late Pliocene and Early Pleistocene dinoflagellate cyst assemblages have been compared with geochemical data from the same samples. Mg/Ca ratios of Globigerina bulloides were measured to estimate the spring-summer sea-surface temperatures from four North Atlantic IODP/DSDP sites. Currently, our Pliocene-Pleistocene database contains 204 dinoflagellate cyst samples calibrated to geochemical data. This palaeo-database is compared with modern North Atlantic and global datasets. The focus lies in the quantitative relationship between Mg/Ca-based (i.e. spring-summer) sea-surface temperature (SSTMg/Ca) and dinoflagellate cyst distributions. In general, extant species are shown to have comparable spring-summer SST ranges in the past and today, demonstrating that our new approach is valid for inferring spring-summer SST ranges for extinct species. For example, Habibacysta tectata represents SSTMg/Ca values between 10° and 15°C when it exceeds 30% of the assemblage, and Invertocysta lacrymosa exceeds 15% when SSTMg/Ca values are between 18.6° and 23.5°C. However, comparing Pliocene and Pleistocene SSTMg/Ca values with present day summer values for the extant Impagidinium pallidum suggests a greater tolerance of higher temperatures in the past. This species occupies more than 5% of the assemblage at SSTMg/Ca values of 11.6-17.9°C in the Pliocene and Pleistocene, whereas present day summer SSTs are around -1.7 to 6.9°C. This observation questions the value of Impagidinium pallidum as reliable indicator of cold waters in older deposits, and may explain its bipolar distribution.

Chemistry of minerals from amphibole-chlorite veins of ODP Site 209-1270

We examined small-scale shear zones in drillcore samples of abyssal peridotites from the Mid-Atlantic Ridge. These shear zones are associated with veins consisting of chlorite + actinolite/tremolite assemblages, with accessory phases zircon and apatite, and they are interpreted as altered plagiogranite melt impregnations, which originate from hydrous partial melting of gabbroic intrusion in an oceanic detachment fault. Ti-in-zircon thermometry yields temperatures around 820°C for the crystallization of the evolved melt. Reaction path modeling indicates that the alteration assemblage includes serpentine of the adjacent altered peridotites. Based on the model results, we propose that formation of chlorite occurred at higher temperatures than serpentinization, thus leading to strain localization around former plagiogranites during alteration. The detachment fault represents a major pathway for fluids through the oceanic crust, as evidenced by extremely low d18O of altered plagiogranite veins (+3.0-4.2 per mil) and adjacent serpentinites (+ 2.6-3.7 per mil). The uniform oxygen isotope data indicate that fluid flow in the detachment fault system affected veins and adjacent host serpentinites likewise.

Soil characteristics and decomposition of organic matter on Anchorage, Signy and Falkland Islands

Antarctic terrestrial ecosystems have poorly developed soils and currently experience one of the greatest rates of climate warming on the globe. We investigated the responsiveness of organic matter decomposition in Maritime Antarctic terrestrial ecosystems to climate change, using two study sites in the Antarctic Peninsula region (Anchorage Island, 67°S; Signy Island, 61°S), and contrasted the responses found with those at the cool temperate Falkland Islands (52°S). Our approach consisted of two complementary methods: (1) Laboratory measurements of decomposition at different temperatures (2, 6 and 10 °C) of plant material and soil organic matter from all three locations. (2) Field measurements at all three locations on the decomposition of soil organic matter, plant material and cellulose, both under natural conditions and under experimental warming (about 0.8 °C) achieved using open top chambers. Higher temperatures led to higher organic matter breakdown in the laboratory studies, indicating that decomposition in Maritime Antarctic terrestrial ecosystems is likely to increase with increasing soil temperatures. However, both laboratory and field studies showed that decomposition was more strongly influenced by local substratum characteristics (especially soil N availability) and plant functional type composition than by large-scale temperature differences. The very small responsiveness of organic matter decomposition in the field (experimental temperature increase <1 °C) compared with the laboratory (experimental increases of 4 or 8 °C) shows that substantial warming is required before significant effects can be detected.

Planktonic foraminifera fluxes and stable isotope ratios in the Irminger Sea

Past water column stratification can be assessed through comparison of the d18O of different planktonic foraminiferal species. The underlying assumption is that different species form their shells simultaneously, but at different depths in the water column. We evaluate this assumption using a sediment trap time-series of Neogloboquadrina pachyderma (s) and Globigerina bulloides from the NW North Atlantic. We determined fluxes, d18O and d13C of shells from two size fractions to assess size-related effects on shell chemistry and to better constrain the underlying causes of isotopic differences between foraminifera in deep-sea sediments. Our data indicate that in the subpolar North Atlantic differences in the seasonality of the shell flux, and not in depth habitat or test size, determine the interspecies Delta d18O. N. pachyderma (s) preferentially forms from early spring to late summer, whereas the flux ofG. bulloides peaks later in the season and is sustained until autumn. Likewise, seasonality influences large and small specimens differently, with large shells settling earlier in the season. The similarity of the seasonal d18O patterns between the two species indicates that they calcify in an overlapping depth zone close to the surface. However, their d13C patterns are markedly different (>1 per mil). Both species have a seasonally variable offset from d13CDIC that appears to be governed primarily by temperature, with larger offsets associated with higher temperatures. The variable offset from d13CDIC implies that seasonality of the flux affects the fossil d13C signal, which has implications for reconstruction of the past oceanic carbon cycle.

Characterization of diagenetic fluids at ODP Site 103-639

Site 639, drilled during Leg 103 of the Ocean Drilling Program, penetrated an Upper Jurassic to Lower Cretaceous carbonate platform on a tilted fault block along the Galicia margin off the northwest Iberian Peninsula. The carbonate platform is composed primarily of a sequence of dolomite overlying limestone. Samples were analyzed for mineral chemistry, stable isotope geochemistry, fluid inclusion microthermometry, and volatile contents and by dolomite pyrolysis mass spectrometry for trace sulfate minerals. The dolomite recovered from the Galicia margin at Site 639 formed during shallow burial from sulfate-bearing, hypersaline brines at slightly elevated temperatures. The light oxygen isotopic signatures of the dolomite are interpreted as the result of the evaporative loop and slightly elevated temperatures during dolomite formation or from reequilibration at higher temperatures during deeper burial. The hypersalinity is interpreted to be associated with a nearby, shallow restricted basin that formed during rifting of the Iberian margin from Newfoundland. The dolomitization of the platform is therefore a by-product of the rifting.

Texture, geochemistry, isotope composition, petrography and epoch at DSDP Site 93-603

Abundant Fe-Mn carbonate concretions (mainly siderite, manganosiderite, and rhodochrosite) were found in the hemipelagic claystones of Site 603 on the eastern North American continental rise. They occur as nodules, micronodules, or carbonate-replaced burrow fills and layers at a subbottom depth of between ~ 120 (Pliocene) and 1160 m (Albian-Cenomanian). In general, the Fe-Mn carbonate concretions form from CO3- produced by the microbiological degradation of organic matter in the presence of abundant Fe + or Mn + and very low S- concentrations. However, there is also some evidence for diagenetic replacement of preexisting calcite by siderite. The carbon isotope composition of diagenetic Fe-Mn carbonate nodules is determined by CO2 reduction during methanogenesis. Carbonate nodules in Cretaceous sediments at sub-bottom depths of 1085 and 1160 m have distinctly lower d13C values (- 12.2 and - 12.9 per mil) than Neogene siderites, associated with abundant biogenic methane in the pore space (-8.9 to 1.7 per mil between 330 and 780 m depth). Since no isotopic zonation could be detected within individual nodules, we assume that the isotopic composition reflects more or less geochemical conditions at the present burial depth of the carbonate nodules. Carbonates did not precipitate within the zone of sulfate reduction (approximately 0.01 to 10 m), where all of the pyrite was formed. The oxygen isotope composition indicates precipitation from seawater-derived interstitial waters. The d18O values decrease with increasing burial depth from + 5.1 to - 1.2 per mil, suggesting successively higher temperatures during carbonate formation.

Geochemistry of volcanic glass from ODP holes 129-801C and 185-801C in the western Pacific

Major element chemistry of basalt from the southern East Pacific Rise (EPR) is different from that of the EPR at the time of the formation of the Pacific Plate at 170 Ma.Glass recovered from Jurassic age (170 Ma) Pacific ocean crust (Bartolini and Larson, 2001, doi:10.1130/0091-7613(2001)029<0735:PMATPS>2.0.CO;2) at Ocean Drilling Program Hole 801C records higher Fe8 (10.77 wt%) and marginally lower Na8 (2.21 wt%) compared to the modern EPR, suggesting deeper melting and a temperature of initial melting that was 60°C hotter than today.Trace element ratios such as La/Sm and Zr/Y, on the other hand, show remarkable similarities to the modern southern EPR, indicating that Site 801 was not generated on a hotspot-influenced ridge and that mantle composition has changed little in the Pacific over the past 170 Ma. Our results are consistent with the observation that mid-ocean ridge basalts (MORBs) older than 80 Ma were derived by higher temperature melting than are modern MORBs (Humler et al., 1999, doi:10.1016/S0012-821X(99)00218-6), which may have been a consequence of the Cretaceous superplume event in the Pacific.Site 801 predates the formation of Pacific oceanic plateaus and 801C basalt chemistry indicates that higher temperatures of mantle melting beneath Pacific ridges preceded the initiation of the superplume.

Calcareous nannofossil assemblages of Ocean Anoxic Event 2 in ODP Sites 207-1258 and 207-1260

The calcareous nannofossils of the Cenomanian/Turonian boundary interval of Sites 1258 and 1260 (Ocean Drilling Program Leg 207) have been studied in order to understand the depositional environment during Oceanic Anoxic Event 2 (OAE2) in the equatorial Atlantic. Nannofossil assemblages show a significant change in relative abundances during the positive d13Corg excursion interval. The strong increase of the high productivity indicator Zeugrhabdotus erectus and the simultaneous decrease of the oligotrophic taxa Watznaueria barnesiae and Watznaueria fossacincta are indicative of enhanced fertility. The decrease of Eprolithus floralis may be attributed to the surface-water temperature increase during OAE2, which is, however, not very significant (~2-3 °C), as suggested by published TEX86 data. It seems more likely that the decrease of E. floralis during OAE2 was evoked by the breakdown of water-column stratification, indicating it as a deep-dwelling species, which prefers stratified waters with a deep nutricline. Prediscosphaera spp. and Retecapsa ficula, which show a significant increase in relative abundances during OAE2, seem to prefer eutrophic environments, while Amphizygus brooksii and Zeugrhabdotus noeliae lower surface-water fertility. Gartnerago segmentatum, Broinsonia spp., Watznaueria biporta, and Seribiscutum gaultensis decrease in abundances during OAE2. It is not clear if they preferred an oligotrophic environment, cooler surface-waters, or if they were inhabitants of the lower photic zone. Published geochemical data suggest that enhanced fertility and higher temperatures during OAE2 may have been caused by submarine volcanic activity through the release of biolimiting micronutrients into the ocean and carbon dioxide into the atmosphere. The breakdown of water-column stratification may have increased further nutrient availability.

Mg/Ca-temperature proxy in benthic foraminifera from the Florida Strait

Over the past decade, the ratio of Mg to Ca in foraminiferal tests has emerged as a valuable paleotemperature proxy. However, large uncertainties remain in the relationships between benthic foraminiferal Mg/Ca and temperature. Mg/Ca was measured in benthic foraminifera from 31 high-quality multicore tops collected in the Florida Straits, spanning a temperature range of 5.8° to 18.6°C. New calibrations are presented for Uvigerina peregrina, Planulina ariminensis, Planulina foveolata, and Hoeglundina elegans. The Mg/Ca values and temperature sensitivities vary among species, but all species exhibit a positive correlation that decreases in slope at higher temperatures. The decrease in the sensitivity of Mg/Ca to temperature may potentially be explained by Mg/Ca suppression at high carbonate ion concentrations. It is suggested that a carbonate ion influence on Mg/Ca may be adjusted for by dividing Mg/Ca by Li/Ca. The Mg/Li ratio displays stronger correlations to temperature, with up to 90% of variance explained, than Mg/Ca alone. These new calibrations are tested on several Last Glacial Maximum (LGM) samples from the Florida Straits. LGM temperatures reconstructed from Mg/Ca and Mg/Li are generally more scattered than core top measurements and may be contaminated by high-Mg overgrowths. The potential for Mg/Ca and Mg/Li as temperature proxies warrants further testing.

Biological traits, and egg and fecal pellet production of Calanus finmarchicus and Calanus glacialis in situ and during experiments

The effects of temperature and food was examined for Calanus finmarchicus and C. glacialis during 3 phases of the phytoplankton spring bloom in Disko Bay, western Greenland. The 2 species were collected during pre-bloom, bloom, and post-bloom and exposed to temperatures from 0 to 10°C, combined with deficient or excess food. Fecal pellet and egg production were measured as indices for grazing and secondary production, respectively. Furthermore, changes in body carbon, nitrogen, and lipid content were measured. C. glacialis sampled before the bloom and incubated with excess food exhibited high specific egg production at temperatures between 0 and 2.5°C. Higher temperatures did not increase egg production considerably, whereas egg production for C. finmarchicus more than tripled between 2.5 and 5°C. Starved C. glacialis produced eggs at all temperatures stimulated by increasing temperatures, whereas starved C. finmarchicus needed temperatures above 5°C to produce eggs fueled by their lipid stores. Few C. finmarchicus had mature gonads at the initiation of the pre-bloom and bloom experiment, and egg production of C. finmarchicus therefore only increased as the ratio of individuals with mature gonads increased. During the bloom, both C. glacialis and C. finmarchicus used the high food availability for egg production, while refueling or exhausting their lipid stores, respectively. Finally, during the post-bloom experiment, production was low by C. finmarchicus, whereas C. glacialis had terminated production. Our results suggest that a future warmer ocean will reduce the advantage of early spawning by C. glacialis and that C. finmarchicus will become increasingly prevalent.

Microcharcoal concentration and elongation, and age model of sediment core MD96-2098

Although grassland and savanna occupy only a quarter of the world's vegetation, burning in these ecosystems accounts for roughly half the global carbon emissions from fire. However, the processes that govern changes in grassland burning are poorly understood, particularly on time scales beyond satellite records. We analyzed microcharcoal, sediments, and geochemistry in a high-resolution marine sediment core off Namibia to identify the processes that have controlled biomass burning in southern African grassland ecosystems under large, multimillennial-scale climate changes. Six fire cycles occurred during the past 170,000 y in southern Africa that correspond both in timing and magnitude to the precessional forcing of north-south shifts in the Intertropical Convergence Zone. Contrary to the conventional expectation that fire increases with higher temperatures and increased drought, we found that wetter and cooler climates cause increased burning in the study region, owing to a shift in rainfall amount and seasonality (and thus vegetation flammability). We also show that charcoal morphology (i.e., the particle's length-to-width ratio) can be used to reconstruct changes in fire activity as well as biome shifts over time. Our results provide essential context for understanding current and future grassland-fire dynamics and their associated carbon emissions.