Clostridium difficile spores must germinate in vivo to become actively growing bacteria in order to produce the toxins that are necessary for disease. C. difficile spores germinate in vitro in ...response to certain bile acids and glycine. In other sporulating bacteria, proteins embedded within the inner membrane of the spore sense the presence of germinants and trigger the release of Ca⁺⁺-dipicolinic acid (Ca⁺⁺-DPA) from the spore core and subsequent hydrolysis of the spore cortex, a specialized peptidoglycan. Based upon homology searches of known germinant receptors from other spore-forming bacteria, C. difficile likely uses unique mechanisms to recognize germinants. Here, we identify the germination-specific protease, CspC, as the C. difficile bile acid germinant receptor and show that bile acid-mediated germination is important for establishing C. difficile disease in the hamster model of infection. These results highlight the importance of bile acids in triggering in vivo germination and provide the first description of a C. difficile spore germinant receptor. Blocking the interaction of bile acids with the C. difficile spore may represent an attractive target for novel therapeutics.
Bacterial spore germination is a process whereby a dormant spore returns to active, vegetative growth, and this process has largely been studied in the model organism Bacillus subtilis. In B. ...subtilis, the initiation of germinant receptor-mediated spore germination is divided into two genetically separable stages. Stage I is characterized by the release of dipicolinic acid (DPA) from the spore core. Stage II is characterized by cortex degradation, and stage II is activated by the DPA released during stage I. Thus, DPA release precedes cortex hydrolysis during B. subtilis spore germination. Here, we investigated the timing of DPA release and cortex hydrolysis during Clostridium difficile spore germination and found that cortex hydrolysis precedes DPA release. Inactivation of either the bile acid germinant receptor, cspC, or the cortex hydrolase, sleC, prevented both cortex hydrolysis and DPA release. Because both cortex hydrolysis and DPA release during C. difficile spore germination are dependent on the presence of the germinant receptor and the cortex hydrolase, the release of DPA from the core may rely on the osmotic swelling of the core upon cortex hydrolysis. These results have implications for the hypothesized glycine receptor and suggest that the initiation of germinant receptor-mediated C. difficile spore germination proceeds through a novel germination pathway.
Clostridium difficile infects antibiotic-treated hosts and spreads between hosts as a dormant spore. In a host, spores germinate to the vegetative form that produces the toxins necessary for disease. C. difficile spore germination is stimulated by certain bile acids and glycine. We recently identified the bile acid germinant receptor as the germination-specific, protease-like CspC. CspC is likely cortex localized, where it can transmit the bile acid signal to the cortex hydrolase, SleC. Due to the differences in location of CspC compared to the Bacillus subtilis germinant receptors, we hypothesized that there are fundamental differences in the germination processes between the model organism and C. difficile. We found that C. difficile spore germination proceeds through a novel pathway.
Infections caused by Clostridium difficile have increased steadily over the past several years. While studies on C. difficile virulence and physiology have been hindered, in the past, by lack of ...genetic approaches and suitable animal models, newly developed technologies and animal models allow these processes to be studied in detail. One such advance is the generation of a mouse-model of C. difficile infection. The development of this system is a major step forward in analyzing the genetic requirements for colonization and infection. While important, it is equally as important in understanding what differences exist between mice and humans. One of these differences is the natural bile acid composition. Bile acid-mediated spore germination is an important step in C. difficile colonization. Mice produce several different bile acids that are not found in humans. These muricholic acids have the potential to impact C. difficile spore germination. Here we find that the three muricholic acids (α-muricholic acid, β-muricholic acid and ω-muricholic acid) inhibit C. difficile spore germination and can impact the growth of vegetative cells. These results highlight an important difference between humans and mice and may have an impact on C. difficile virulence in the mouse-model of C. difficile infection.
To better understand the evolutionary history of the African continental crust, a combined U–Pb, Lu–Hf and O isotopic study has been carried out by in situ analyses of approximately 450 detrital ...zircon grains from the Niger, Nile, Congo, Zambezi and Orange Rivers. The U–Pb isotopic data show age peaks at ca. 2.7, 2.1–1.8, 1.2–1.0, ca. 0.8, 0.7–0.5 and ca. 0.3Ga. These peaks, with the exception of the one at ca. 0.8Ga, correspond with the assembly of supercontinents. Furthermore, the detrital zircons that crystallized during these periods of supercontinent assembly have dominantly non-mantle-like O and Hf isotopic signatures, in contrast to the ca. 0.8Ga detrital zircons which have juvenile characteristics. These data can be interpreted as showing that continental collisions during supercontinent assembly resulted in supermountain building accompanied by remelting of older continental crust, which in turn led to significant erosion of young igneous rocks with non-mantle-like isotopic signatures. Alternatively, the data may indicate that the major mode of crustal development changed during the supercontinent cycle: the generation of juvenile crust in extensional settings was dominant during supercontinent fragmentation, whereas the stabilization of the generated crust via crustal accretion and reworking was important during supercontinent assembly. The Lu–Hf and O isotope systematics indicate that terreigneous sediments could attain elevated 18O/16O via prolonged sediment–sediment recycling over long crustal residence time, and also that reworking of carbonate and chert which generally have elevated 18O/16O and low Hf contents is minor in granitoid magmatism. The highest 18O/16O in detrital zircon abruptly increased at ca. 2.1Ga and became nearly constant thereafter. This indicates that reworking of mature sediments increased abruptly at that time, probably as a result of a transition in the dynamics of either granitoid crust formation or sedimentary evolution. To estimate the mantle-extraction age of the reworked crust, we have calculated arc mantle Hf model ages for the detrital zircons using O isotopic data to constrain the Lu/Hf used in the model age calculation. The Hf model age histograms for each period of detrital zircons suggest that a significant amount of the African continental crust was generated in the Paleo-Mesoproterozoic likely by mafic magmatism, and subsequently reworked into younger granitoid crust with varying crustal residence times.
Chronic exposure to toxic metals is a serious global health concern. However, population-wide biomonitoring is costly and carries several sampling constraints. Though hair sampling can be a useful ...way to assess environmental exposure, external contamination is a long-standing concern, and a pre-cleaning step prior to metal quantification has long been recommended despite a lack of evidence for its efficacy. In this study, we quantified the spatial distribution of 16 elements in unwashed human hair samples using Laser Ablation Inductively Coupled Plasma Mass Spectrometry (LA-ICP-MS), then tested how two common pre-cleaning treatments (Triton-ethanol, Triton-nitric acid) affected metal content in external and interior layers of hair using LA-ICP-MS. We show that elements differ in their spatial distribution across hair and that pre-cleaning is not consistent in its effect on element concentrations and decreases interior concentrations of some elements. We demonstrate that differences among individuals can be quantified reliably with LA-ICP-MS analysis of interior concentrations of unwashed hair. Our study tests the widespread notion that pre-cleaning is essential in analyses of hair for environmental exposure to metals, and examines the benefits of a unified approach to analysis of metals in hair using LA-ICP-MS.
Cysteine‐reactive small molecules are used as chemical probes of biological systems and as medicines. Identifying high‐quality covalent ligands requires comprehensive kinetic analysis to distinguish ...selective binders from pan‐reactive compounds. Quantitative irreversible tethering (qIT), a general method for screening cysteine‐reactive small molecules based upon the maximization of kinetic selectivity, is described. This method was applied prospectively to discover covalent fragments that target the clinically important cell cycle regulator Cdk2. Crystal structures of the inhibitor complexes validate the approach and guide further optimization. The power of this technique is highlighted by the identification of a Cdk2‐selective allosteric (type IV) kinase inhibitor whose novel mode‐of‐action could be exploited therapeutically.
A simple high‐throughput fluorescence assay was developed which rapidly identifies high‐affinity covalent fragments by maximizing kinetic selectivity of protein modification. This platform allows electrophiles with different warheads and reactivities to be simultaneously screened. Using this technology, a new class of allosteric Cdk2 inhibitors was discovered which show a unique selectivity profile and have potential therapeutic use.
We use 369 individual U–Pb zircon ages from 14 granitoid samples collected on five islands in the Cyclades in the Aegean Sea, Greece, for constraining the crystallisation history of I- and S-type ...plutons above the retreating Hellenic subduction zone. Miocene magmatism in the Cyclades extended over a time span from 17 to 11 Ma. The ages for S-type granites are systematically ~2 million years older than those for I-type granites. Considering plutons individually, the zircon data define age spectra ranging from simple and unimodal to complex and multimodal. Seven of the 14 investigated samples yield more than one distinct zircon crystallisation age, with one I-type granodiorite sample from Mykonos Island representing the most complex case with three resolvable age peaks. Two samples from S-type granites on Ikaria appear to have crystallised zircon over 2–3 million years, whereas for the majority of individual samples with multiple zircon age populations the calculated ages deviate by 1–1.5 million years. We interpret our age data to reflect a protracted history involving initial partial melting at deeper lithospheric levels, followed by crystallisation and cooling at shallower crustal levels. Our study corroborates published research arguing that pluton construction is due to incremental emplacement of multiple magma pulses over a few million years. Assuming that multiple age peaks of our 14 samples can indeed serve to quantify time spans for magmatic emplacement, our data suggest that Aegean plutons were constructed over a few million years. Our tectonic interpretation of the U–Pb ages is that the S-type granites resulted from partial melting and migmatisation of the lower crust, possibly starting at ~23 Ma. The I-type granites and associated mafic melts are interpreted to reflect the magmatic arc stage in the Cyclades starting at ~15 Ma.
New U-Pb detrital zircon ages from sandstones have been determined for the Walloon Coal Measures-Springbok Sandstone section of the Surat Basin. In the late Jurassic Surat Basin, there is a shift ...from relatively thick coal measures – the Walloon Coal Measures – consisting of laterally discontinuous coal-seam horizons intercalated with lithic sandstones and mudstones to an overlying coal-poor sandstone-dominated unit – the Springbok Sandstone – that is more laterally continuous. Across the basin, the contact between the two units varies from transitional to unconformable, and the sandstone becomes less lithic. The nature of the contact and the compositional variability of the sandstone across the basin have implications for predicting communication between gas production fields and potential aquifers in the Walloon Coal Measures and overlying Springbok Sandstone. This study tests whether detrital zircon age (LA-ICP-MS) populations from sandstones within these two units can identify provenance shifts and potential tectonic drivers for basin infill at this time. Detrital zircon ages of both units, informed by paleocurrent measurements showing a southerly flowing depositional system, suggest sediment was sourced from three distinct terranes: the Thomson Orogen in northeastern Queensland; Permian-Triassic granitic intrusions emplaced within the northeast New England Orogen during the Hunter-Bowen Orogeny accretion event; and an inferred but now eroded Jurassic magmatic-arc located along the eastern margin of Gondwana. Two major source-delivery systems, named here Western and Eastern Source systems, are recognized by distinct detrital zircon age populations in both formations in the northern Surat Basin. The Western Source of the Walloon Coal Measures delivered substantial Jurassic age zircons, with minor Pre-Cambrian input; the Eastern Source delivered substantial reworked Precambrian and lesser Jurassic zircons. In contrast, the Western Source of the Springbok Sandstone delivered substantial Jurassic and distinct Triassic zircons dated between ca 234 Ma and 224 Ma in the north-west, while the Eastern Source delivered more pronounced Pre-Cambrian input, with minor Jurassic and absent or minor age zircons in the northeast. Maximum depositional ages for the Springbok Sandstone are approximately 2 Ma younger in the northwestern than in the northeastern Surat Basin. The westward younging could be due to migration of the system and/or differential movement on the bounding faults.
•New insights into spatial and lateral provenance shift between the Middle-Late Jurassic strata, northern Surat Basin.•Multiple fluvial depocenters identified in the northern Surat Basin using U-Pb detrital zircon geochronology.•Potential reactivation of fault system may have caused a provenance shift between the Middle-Late Jurassic strata in Surat Basin•The detrital age populations when combined with MDAs and published CA-TIMS reveal a younging trend from east to west.
The shoshonitic intrusions of eastern Tibet, which range in age from 33 to 41 Ma and in composition from ultramafic (SiO
2
= 42 %) to felsic (SiO
2
= 74 %), were produced during the collision of ...India with Eurasia. The mafic and ultramafic members of the suite are characterized by phenocrysts of phlogopite, olivine and clinopyroxene, low SiO
2
, high MgO and Mg/Fe ratios, and olivine forsterite contents of Fo
87
to Fo
93
, indicative of equilibrium with mantle olivine and orthopyroxene. Direct melting of the mantle, on the other hand, could not have produced the felsic members. They have a phenocryst assemblage of plagioclase, amphibole and quartz, high SiO
2
and low MgO, with Mg/Fe ratios well below the values expected for a melt in equilibrium with the mantle. Furthermore, the lack of decrease in Cr with increasing SiO
2
and decreasing MgO from ultramafic to felsic rocks precludes the possibility that the felsic members were derived by fractional crystallization from the mafic members. Similarly, magma mixing, crustal contamination and crystal accumulation can be excluded as important processes. Yet all members of the suite share similar incompatible element and radiogenic isotope ratios, which suggests a common origin and source. We propose that melting for all members of the shoshonite suite was initiated in continental crust that was thrust into the upper mantle at various points along the transpressional Red River-Ailao Shan-Batang-Lijiang fault system. The melt formed by high-degree, fluid-absent melting reactions at high-T and high-P and at the expense of biotite and phengite. The melts acquired their high concentrations of incompatible elements as a consequence of the complete dissolution of pre-existing accessory minerals. The melts produced were quartz-saturated and reacted with the overlying mantle to produce garnet and pyroxene during their ascent. The felsic magmas reacted little with the adjacent mantle and preserved the essential features of their original chemistry, including their high SiO
2
, low Ni, Cr and MgO contents, and low Mg/Fe ratio, whereas the mafic and ultramafic magmas are the result of extensive reaction with the mantle. Although the mafic magmas preserved the incompatible element and radiogenic isotope ratios of their crustal source, buffering by olivine and orthopyroxene extensively modified their MgO, Ni, Cr, SiO
2
contents and Mg/Fe ratio to values dictated by equilibrium with the mantle.
The last major episode of cordilleran‐style tectonism in eastern Australia was the late Paleozoic‐early Mesozoic Gondwanide Orogeny. When exactly this deformation commenced and what caused this phase ...of orogenesis is still debated. Using previous stratigraphic and sedimentological data from pre‐orogenic to syn‐orogenic strata, integrated with new detrital zircon U‐Pb geochronology, we investigated the onset of the Gondwanide Orogeny in the northern New England Orogen (Gogango Overfolded Zone, eastern Australia). The lowermost syn‐orogenic strata display evidence of rapid base‐level change coeval with increasingly abundant and more proximal mass‐wasting deposits, which reflect a shift to static loading‐induced subsidence and the establishment of a ∼1,600‐km long foreland basin system. The syn‐orogenic strata contain abundant detritus sourced from syn‐depositional magmatism, alongside an up‐section increase in detritus derived from uplifted older rocks of the New England Orogen. Detrital zircons show maximum depositional age of ∼276 Ma from immediately above the sequence boundary, and ∼269 Ma from an overlying formation. These results confirm previous suggestions that tectonic forcing of the orogenic hinterland affected subsidence and sedimentation patterns in the easternmost part of the basin during the middle Permian, >11 Myr prior to deformation in the western part of the foreland basin. The onset of the Gondwanide Orogeny in eastern Australia, and in other sectors of the Gondwanan margin, likely resulted from a tectonic switch from crustal extension to contraction, which was driven by increased convergence rates due to a plate reorganization event following the final assembly of Pangea.
Key Points
New detrital zircon U‐Pb data and reinterpreted stratigraphic and sedimentological data unravel the onset of Gondwanide Orogeny in eastern Australia
In the northern New England Orogen, orogenesis commenced at ∼276 Ma
Orogenesis was likely driven by increased convergence rates due to a plate reorganization event following the final assembly of Pangea
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