Étude biostratigraphique et paléoenvironnementale des diatomées pyritisées de l'intervalle Paléocène supérieur-Éocène inférieur du sondage de Saint-Josse (France)

This paper presents the results of a biostratigraphic analysis of pyritized diatom assemblages in the Saint-Josse core (CC82) located in the northern part of the Paris basin (coordinates: X =555?000, Y =307?324 and Z =56.9 m). During the Paleocene-Eocene this locality was situated in a small bight of the North Sea Basin. The results are obtained from sediments of the Thanet and Mont-Bernon group equivalents belonging to the Paleocene-Eocene interval. Thirteen samples were studied and eleven contain pyritized diatoms. By epigenesis, the siliceous frustule of the diatom is changed into a pyritized skeleton with all the ornamentation very well preserved such as the velum, the areolae, the labiate processes, etc. Dispersive X-Ray analyses show this complete replacement of silica by pyrite. Another form of pyritization is crystallization of pyrite (in cubes, pyritohedrons, octahedrons, framboids) in internal cavities of the diatoms to produce internal moulds, also called steinkerns, which preserve the external morphology of the diatoms and some details such as the girdle. Despite pyritization, diatoms can often be recognized at the species level; brackish and marine species were observed but no fresh water specimens were encountered. Pyrite is easily altered into iron oxides. In outcrops, pyritized diatoms can be obscured or destroyed by oxidation. We have chosen borehole samples for this study to avoid the affects of weathering. Deposits from the Sorrus Unit (of the Thanet Group) have low abundances of pyritized diatoms (3 to 25 specimens per sample). This unit contains allochtonous diatom assemblages comprising Actinoptychussenarius, Coscinodiscus morsianus var. morsianus, and Coscinodiscus morsianus var. moelleri. Sample 34.31-34.45 from this unit is particularly interesting. It contains three specimens of Paralia siberica var. laevis, which is typical of an estuarine or littoral paleoenvironment, and corresponds to the more continental deposit of the Sorrus Unit. Seven samples, taken in the Argile de Saint-Aubin Unit (from the Mont-Bernon Group), contain more than 200 pyritized diatoms and between 9 to 16 different species per sample. A brackish assemblage (Actinoptychus senarius, Coscinodiscus commutatus and Stellarima microtrias) and a marine assemblage (C. morsianus var. morsianus, Coscinodiscus var. moelleri, Fenestrella antiqua, Odontellaheibergii, Rhizosolenia sp. 1 and Trinacria regina) are observed. The brackish assemblage expands in the upper part of the Saint-Aubin Unit indicating more widespread brackish paleoenvironmental conditions. Pyritized diatoms are important fossil markers in the Paris basin mainly when siliceous fossils (silicoflagellates or radiolarians) and others microfossils (foraminifers and dinoflagellates) are very poor or absent. Two diatom assemblages have been identified in the Saint-Josse borehole. The first assemblage D1, comprising C. morsianus var. moelleri (high occurrence), C. morsianus (a few) and T. regina (rare), is recorded in the Sorrus, La Calotterie, Bois Gorguette, Le Goulet and lower Saint-Aubin Units. The second assemblage D2, characterized by great abundance of F. antiqua and decreasing numbers of C. morsianus species, is found in the upper part of the Saint-Aubin Unit. These diatom assemblages can be correlated with other biozonations established in the North Sea basin and based on nannoplankton and diatoms (King, 1983), diatoms (Mitlehner, 1996) and calcareous nannoplankton (Martini, 1971). F. antiqua and C. morsianus var. moelleri are the more important diatom species encountered. F. antiqua (previously named Coscinodiscus sp. 1, King, 1983) is the fossil marker used by King to define his NSP4 biozone. The base of NSP4 corresponds to the lowest occurrence of F. antiqua and the top is marked by the highest occurrence of this species. The Sorrus, La Calotterie, Bois Gorguette, Le Goulet and Saint-Aubin (“Sparnacian” stage) Units, the last one comprising F. antiqua (increasing to acme), correspond to the NSP4 zone of King (1983). The Mont-Hulin and Château de la Bruyère Units correspond probably to the NSP5 zone of King (1983). F. antiqua and C. morsianus var. moelleri were also used by Mitlehner (1996) to differentiate the NSP4a zone (low abundance of F. antiqua and abundance of C. morsianus var. moelleri) from the NSP4b zone (great abundance and acme of F. antiqua). The abundance of C. morsianus var. moelleri is recorded in the lower part of Saint-Aubin Unit and the abundance of F. antiqua (high peak at 46% in the sample 17.87) is found in the upper part of Saint-Aubin Unit. Therefore, Sorrus, La Calotterie, Bois Gorguette, Le Goulet and the lower part of Saint-Aubin Units are correlated with the NSP4a zone and the upper part of Saint-Aubin Unit with the NSP4b zone. By correlating the calcareous nannoplankton biozonation (NP) of Martini (1971) with the biozonation of King (1983), adapted by Mitlehner (1996), the boundary between the NP9 and NP10 zones could be located in the 18.75-17.87 interval and the NP10-NP11 boundary at the top of Saint-Aubin Unit. Lithostratigraphic correlations between the units of the Saint-Josse borehole (Paris basin) and the formations of the North Sea basin can be established. The great abundance of C. morsianus var. moelleri and the great abundance (including acme) of F. antiqua are respectively recorded in the Sele and Balder Formations. Therefore, the interval Sorrus/lower Saint-Aubin Units is correlated with the Sele Formation and upper part of Saint-Aubin Unit with the Balder Formation.     

In vitro evaluation of native entomopathogenic fungi and neem (Azadiractha indica) extracts on Spodoptera frugiperda

The control of Spodoptera frugiperda is based on synthetic insecticides, so some alternatives are the use of entomopathogenic fungi (EF) and neem extract. The objective of the study was to evaluate in vitro effectiveness of native EF and neem extracts on S. frugiperda larvae. Six EF were identified by DNA sequencing of ITS regions from three EF (Fusarium solani, Metarrhizium robertsii, Nigrospora spherica and Penicillium citrinum). They were evaluated in concentrations of 1 × 10⁸ spores/ mL. In addition, a second bioassay was carried out evaluating only F. solani, M. robertsii and N. sphaerica and the addition of vegetable oil. On the other hand, extraction of secondary metabolites from neem seed (Azadirachta indica) was carried out by performing, mass (g) and solvent volume (mL ethanol and water) combinations, which were subjected to microwaves and ultrasound. Subsequently, these extracts were evaluated in concentrations of 3%, 4% and 5%. A survival analysis was performed for each of the bioassays. With respect to the results of the first bioassay, F. solani obtained a probability of survival of 0.476 on the seventh day, while in the second bioassay, M. robertsii obtained 0.488 survival probability. This suggests that the expected percentage of larvae that stay alive on the sixth day is 48.8%. However, in the evaluation of the neem extract the combination 1:12/70% to 4% caused 84% mortality of larvae. The use of native HE and neem extracts has potential for the control of S. frugiperda.     

An Ustilago maydis Mutant Partially Blocked in P45014DM Activity Is Hypersensitive to Azole Fungicides

An Ustilago maydis ergosterol biosynthesis mutant (A14) which is partially blocked in sterol 14alpha-demethylase (P45014DM) activity is described. This mutant accumulated the abnormal 14alpha-methyl sterols, eburicol, 14alpha-methylfecosterol, and obtusifoliol, along with significant amounts of ergosterol. Although the A14 mutant grew nearly as well as the wild type, it was impaired in cell extension growth, which indicated a dysfunction in apical cell wall synthesis. The mutant was also found to be hypersensitive to the azole fungicides penconazole and tebuconazole.     

Influence of Temperature Stress on in Vitro Fertilization and Heat Shock Protein Synthesis in Maize (Zea mays L.) Reproductive Tissues

This study was conducted to investigate the response of maize (Zea mays) male and female mature reproductive tissues to temperature stress. We have tested the fertilization abilities of the stressed spikelets and pollen using in vitro pollination-fertilization to determine their respective tolerance to stress. The synthesis of heat shock proteins (HSPs) was also analyzed in male and female tissues using electrophoresis of ³⁵S-labeled proteins and fluorography, to establish a relationship between the physiological and molecular responses. Pollen, spikelets, and pollinated spikelets were exposed to selected temperatures (4, 28, 32, 36, or 40°C) and tested using an in vitro fertilization system. The fertilization rate is highly reduced when pollinated spikelets are exposed to temperatures over 36°C. When pollen and spikelets are exposed separately to temperature stress, the female tissues appear resistant to 4 hours of cold stress (4°C) or heat stress (40°C). Under heat shock conditions, the synthesis of a typical set of HSPs is induced in the female tissues. In contrast, the mature pollen is sensitive to heat stress and is responsible for the failure of fertilization at high temperatures. At the molecular level, no heat shock response is detected in the mature pollen.     

Hexazonium pararosaniline as a fixative for animal tissues

Hexazonium pararosaniline is a valuable reagent that has been used in enzyme activity histochemistry for 50 years. It is an aqueous solution containing the tris-diazonium ion derived from pararosaniline, an aminotriarylmethane dye, and it contains an excess of nitrous acid that was not consumed in the diazotization reaction. Other investigators have found that immersion for 2 min in an acidic (pH 3.5) 0.0015 M hexazonium pararosaniline solution can protect cryostat sections of unfixed animal tissues from the deleterious effects of aqueous reagents such as buffered solutions used in immunohistochemistry, while preserving specific affinities for antibodies. In the present investigation hexazonium pararosaniline protected lymphoid tissue and striated muscle against the damaging effects of water or saline. The same protection was conferred on unfixed sections treated with dilute nitrous or hydrochloric acid in concentrations similar to those in hexazonium pararosaniline solutions. Model tissues (solutions, gels or films containing gelatin and/or bovine albumin) responded predictably to well known cross-linking (formaldehyde) or coagulant (mercuric chloride) fixatives. Hexazonium pararosaniline solutions prevented the dissolution of protein gels in water only after 9 or more days of contact, during which time considerable swelling occurred. It is concluded that there is no evidence for a “fixative” action of hexazonium pararosaniline. The protective effect on frozen sections of unfixed tissue is attributable probably to the low pH of the solution.     

Viral fusion peptides and identification of membrane-interacting segments

Viral envelope glycoproteins promote infection by mediating fusion between viral and cellular membranes. Fusion occurs after dramatic conformational changes within fusion proteins, leading to the exposure of a short stretch of mostly apolar residues, termed the fusion peptide, which is presumed to insert into the membrane and initiate the fusion process. The typical global composition of fusion peptides, rich in hydrophobic but also in small amino acids such as alanine and glycine, was used here as bait to detect other peptidic segments that can insert into membranes. We so evidenced a similar composition in several cytotoxic peptides, which promote pore formation such as peptides involved in amyloidoses and hydrophobic alpha-hairpins of pore-forming toxins. It is suggested that the structural plasticity observed for several membrane active peptides can be conferred by this particular global amino acid composition, which could be thus used to predict such functional behavior from genome data.     

Lipase and Esterase Activities of Propionibacterium freudenreichii subsp. freudenreichii

The lipase and esterase activities of eight strains of dairy Propionibacterium freudenreichii subsp. freudenreichii were studied. A lipase activity was detected on whole cells and in the culture supernatant. The highest activity was expressed at 45°C and pH 6.8. An esterase activity was also detected in the culture medium. The electrophoresis of the intracellular fractions of the cells revealed from three to six different esterase activities. Two esterases were common to all the strains. The substrate specificity was dependent on each esterase, but no activity was revealed, in our experimental conditions, on ester substrates with a chain length longer than that of butyrate.     

American and korean ginseng tissue cultures: Growth,chemical analysis,and plantlet production

Summary Roots, stems, or leaves of American (Panax quinquefolium) and Korean (Panax ginsing) ginseng were grown as callus or supension tissue cultures. Tissue cultures ofP. ginseng would occasionally form plantlets. The fundamental chemical composition, inorganic analysis, and saponin (panaquilin) content of American and Korean ginseng plants and tissue cultures were determined. The crude saponin content is very similar to, but approximately one-half (1.3%, fresh weight) of that present in ginseng roots. Two-dimensional thin layer chromatographic analysis revealed minor differences in the panaquilins present in American and Korean ginseng tissue cultures. The sapogenin, panaxadiol, was isolated from Korean ginseng callus.     

Possible Links Between Stress Defense and the Tricarboxylic Acid (TCA) Cycle in Francisella Pathogenesis

Francisella tularensis is a highly infectious bacterium causing the zoonotic disease tularemia. In vivo, this facultative intracellular bacterium survives and replicates mainly in the cytoplasm of infected cells. We have recently identified a genetic locus, designated moxR that is important for stress resistance and intramacrophage survival of F. tularensis. In the present work, we used tandem affinity purification coupled to mass spectrometry to identify in vivo interacting partners of three proteins encoded by this locus: the MoxR-like ATPase (FTL_0200), and two proteins containing motifs predicted to be involved in protein–protein interactions, bearing von Willebrand A (FTL_0201) and tetratricopeptide (FTL_0205) motifs. The three proteins were designated here for simplification, MoxR, VWA1, and TPR1, respectively. MoxR interacted with 31 proteins, including various enzymes. VWA1 interacted with fewer proteins, but these included the E2 component of 2-oxoglutarate dehydrogenase and TPR1. The protein TPR1 interacted with one hundred proteins, including the E1 and E2 subunits of both oxoglutarate and pyruvate dehydrogenase enzyme complexes, and their common E3 subunit. Remarkably, chromosomal deletion of either moxR or tpr1 impaired pyruvate dehydrogenase and oxoglutarate dehydrogenase activities, supporting the hypothesis of a functional role for the interaction of MoxR and TPR1 with these complexes. Altogether, this work highlights possible links between stress resistance and metabolism in F. tularensis virulence.Francisella tularensis is responsible for the disease tularamia in a large number of animal species. This highly infectious bacterial pathogen can be transmitted to humans in numerous ways (1, 2, 3), including direct contact with sick animals, inhalation, ingestion of contaminated water or food, or by bites from ticks, mosquitoes, or flies. Four different subspecies (subsp.) of F. tularensis that differ in virulence and geographic distribution exist, designated subsp. tularensis (type A), subsp. holarctica (type B), subsp. Novicida, and subsp. mediasiatica, respectively. F. tularensis subsp. tularensis is the most virulent subspecies causing a severe disease in humans, whereas F. tularensis subsp. holarctica causes a similar disease but of less severity (4). Because of its high infectivity and lethality, F. tularensis is considered a potential bioterrorism agent (5).F. tularensis is able to survive and to replicate in the cytoplasm of a variety of infected cells, including macrophages. To resist this stressful environment, the bacterium must have developed stress resistance mechanisms, most of which are not yet well characterized. We recently reported the identification of a novel genetic locus that is important for stress resistance and intracellular survival of F. tularensis (6). This locus was designated moxR because the first gene FTL_0200, encodes a protein belonging to the AAA+ ATPase of the MoxR family ((7) and references therein). The data obtained in that first study had led us to suggest that the F. tularensis MoxR-like protein might constitute, in combination with other proteins of the locus, a chaperone complex contributing to F. tularensis pathogenesis.To further validate this hypothesis and expand our initial observations, we here decided to perform tandem affinity purification (TAP),¹ using a dual affinity tag approach coupled to mass spectroscopy analyses (8), to identify proteins interacting in vivo with three proteins encoded by the proximal portion of the moxR locus. For this, we chose as baits: the MoxR-like protein (FTL_0200) and two proteins bearing distinct motifs possibly involved in protein–protein interactions, FTL_0201 (Von Willebrand Factor Type A domain, or VWA) and FTL_0205 (tetratrichopeptide repeat or TPR). The three proteins were designated here for simplification, MoxR, VWA1, and TPR1; and the corresponding genes moxR, vwa1, and tpr1, respectively.VWA domains are present in all three kingdoms of life. They consist of a β-sheet sandwiched by multiple α helices. Frequently, VWA domain-containing proteins function in multiprotein complexes (9). TPR typically contain 34 amino acids. Many three-dimensional structures of TPR domains have been solved, revealing amphipathic helical structures (10). TPR-containing proteins are also found in all kingdoms of life. They can be involved in a variety of functions, and generally mediate protein–protein interactions. In the past few years, several TPR-related proteins have been shown to be involved in virulence mechanisms in pathogenic bacteria ((11) and references therein).Our proteomic approach allowed us to identify a series of protein interactants for each of the three moxR-encoded proteins. Remarkably, the protein TPR1 interacted with all the subunits of the pyruvate dehydrogenase (PDH) and 2-oxoglutarate dehydrogenase (OGDH) complexes. Furthermore, inactivation of tpr1 also severely impaired the activities of these two enzymes. Inactivation of tpr1 affected bacterial resistance to several stresses (and in particular oxidative stress), intramacrophagic bacterial multiplication and bacterial virulence in the mouse model. Functional implications and possible relationship between bacterial metabolism, stress defense, and bacterial virulence are discussed.