José L. Moreno

Effects of a cadmium-contaminated sewage sludge compost on dynamics of organic matter and microbial activity in an arid soil

Abstract An incubation experiment lasting 120 days was carried out to ascertain the effect on the soil microbial activity and organic matter mineralization of adding a sewage sludge compost contaminated with two different levels of Cd to an arid soil. Two composts, with a low (2 mg kg–1) and high (815 mg kg–1) Cd content, respectively, were used in this experiment. Both composts increased the total organic C, humic substance and water-soluble C contents, the beneficial effects still being noticeable after 120 days of incubation. The most labile C fraction (water-soluble C) was the most sensitive to the high Cd content. The high Cd concentration decreased soil microbial biomass C and stimulated the metabolic activity of the microbial biomass, the metabolic quotient (qCO2) revealing itself to be a very sensitive index of the stress that the incorporation of a Cd-contaminated sewage sludge compost causes in a soil. The effect of Cd contamination on enzyme activities (urease, protease that hydrolyse N-α-benzoil-l-arginamide, phosphatase, and β-glucosidase) depended on the enzyme studied.

Construction of a Severe Acute Respiratory Syndrome Coronavirus Infectious cDNA Clone and a Replicon To Study Coronavirus RNA Synthesis

ABSTRACT The engineering of a full-length infectious cDNA clone and a functional replicon of the severe acute respiratory syndrome coronavirus (SARS-CoV) Urbani strain as bacterial artificial chromosomes (BACs) is described in this study. In this system, the viral RNA was expressed in the cell nucleus under the control of the cytomegalovirus promoter and further amplified in the cytoplasm by the viral replicase. Both the infectious clone and the replicon were fully stable in Escherichia coli. Using the SARS-CoV replicon, we have shown that the recently described RNA-processing enzymes exoribonuclease, endoribonuclease, and 2′-O-ribose methyltransferase were essential for efficient coronavirus RNA synthesis. The SARS reverse genetic system developed as a BAC constitutes a useful tool for the study of fundamental viral processes and also for developing genetically defined vaccines.

Transference of heavy metals from a calcareous soil amended with sewage-sludge compost to barley plants

Barley plants were cultivated in a calcareous soil (pH 8.77) amended, at different rates, with sewage-sludge composts containing different heavy-metal contents and the transference of these heavy metals to the plant was studied. The addition to the soil of contaminated compost (mainly in Cd) at a high rate led to a decrease in grain yield but did not affect straw yield. Cadmium and Zn were easily absorbed by barley plants, increasing their concentration with respect to the control in plants grown in the soil amended with compost containing high amounts of these metals. However, Ni and particularly Cu were retained by organic matter and were not transferred to plants. Regardless of the compost heavy-metal contents, plants grown in amended soils showed higher N and P contents than control plants. The concentrations of Cd and Zn in soils were positively correlated with Cd and Zn contents in the plants, while no correlation was found between the concentrations of Cu or Ni in soil and plant. After cultivation, amended soils showed a better nutritional state than control soil (higher N-NO3− and total- and available-P than the control).

Persistence of immobilised and total urease and phosphatase activities in a soil amended with organic wastes

This paper reports on the persistence of total and immobilised enzyme activities (urease and phosphatase) in a soil amended with organic wastes containing high levels of total-urease and phosphatase activity. Fresh organic materials showed the highest values for both total-enzymatic activities. The addition of organic waste to soil increased both total-enzymatic activities in the soil, which, after 360 days, showed values above those of the control. Immobilised enzymes were also higher in the fresh wastes than in the soil with compost, while the specific enzymatic activity levels (enzymatic activity per unit of carbon) were similar. The immobilised urease activity was greater in the amended soil than in the control. At the beginning of the incubation period, the immobilised urease activity was significantly higher in the soil amended with fresh organic wastes than with compost. However, this activity decreased with incubation, whilst the compost-immobilised urease activity increased with time. The effect of organic amendment on immobilised phosphatase activity was similar to that shown by immobilised urease but less pronounced. The persistence of both enzymes was significantly higher in the soil amended with compost than in that amended with fresh materials.

Coronavirus nucleocapsid protein is an RNA chaperone.

Abstract RNA chaperones are nonspecific nucleic acid binding proteins with long disordered regions that help RNA molecules to adopt its functional conformation. Coronavirus nucleoproteins (N) are nonspecific RNA-binding proteins with long disordered regions. Therefore, we investigated whether transmissible gastroenteritis coronavirus (TGEV) N protein was an RNA chaperone. Purified N protein enhanced hammerhead ribozyme self-cleavage and nucleic acids annealing, which are properties that define RNA chaperones. In contrast, another RNA-binding protein, PTB, did not show these activities. N protein chaperone activity was blocked by specific monoclonal antibodies. Therefore, it was concluded that TGEV N protein is an RNA chaperone. In addition, we have shown that purified severe acute respiratory syndrome (SARS)-CoV N protein also has RNA chaperone activity. In silico predictions of disordered domains showed a similar pattern for all coronavirus N proteins evaluated. Altogether, these data led us to suggest that all coronavirus N proteins might be RNA chaperones.

Application of composted sewage sludges contaminated with heavy metals to an agricultural soil

Abstract To elucidate the mechanism of transfer of heavy metals into the food chain, an experiment was carried out with a calcareous soil, to which two different doses of a sewage sludge compost contaminated with either Cd or Zn, Cd, Cu, and Ni were applied. A crop of lettuce was then grown in the amended soils. The application of sewage sludge composts to a calcareous soil lowered the soils pH, although the value was always around 8 at the end of the experiment. Electric conductivity rose with organic amendment. As anticipated, such an amendment improved the nutritional level of the soils, particularly Nand P, both total and available. Plant yields were negatively affected by organic amendments contaminated with heavy metals, the most dangerous in our experiment being Cd and Zn since this metals easily taken up by plants. As Ni and Cu form insoluble complexes with the organic matter of the sewage sludge composts they are not readily absorbed. Of the metals studied, Cd and Zn showed the highest bioavaila...

Role of Nucleotides Immediately Flanking the Transcription-Regulating Sequence Core in Coronavirus Subgenomic mRNA Synthesis

ABSTRACT The generation of subgenomic mRNAs in coronavirus involves a discontinuous mechanism of transcription by which the common leader sequence, derived from the genome 5′ terminus, is fused to the 5′ end of the mRNA coding sequence (body). Transcription-regulating sequences (TRSs) precede each gene and include a conserved core sequence (CS) surrounded by relatively variable sequences (5′ TRS and 3′ TRS). Regulation of transcription in coronaviruses has been studied by reverse-genetics analysis of the sequences immediately flanking a unique CS in the Transmissible gastroenteritis virus genome (CS-S2), located inside the S gene, that does not lead to detectable amounts of the corresponding mRNA, in spite of its canonical sequence. The transcriptional inactivity of CS-S2 was genome position independent. The presence of a canonical CS was not sufficient to drive transcription, but subgenomic synthesis requires a minimum base pairing between the leader TRS (TRS-L) and the complement of the body TRS (cTRS-B) provided by the CS and its adjacent nucleotides. A good correlation was observed between the free energy of TRS-L and cTRS-B duplex formation and the levels of subgenomic mRNA S2, demonstrating that base pairing between the leader and body beyond the CS is a determinant regulation factor in coronavirus transcription. In TRS mutants with increasing complementarity between TRS-L and cTRS-B, a tendency to reach a plateau in ΔG values was observed, suggesting that a more precise definition of the TRS limits might be proposed, specifically that it consists of the central CS and around 4 nucleotides flanking 5′ and 3′ the CS. Sequences downstream of the CS exert a stronger influence on the template-switching decision according to a model of polymerase strand transfer and template switching during minus-strand synthesis.

OSIRIS observations of meter-sized exposures of H2O ice at the surface of 67P/Churyumov-Gerasimenko and interpretation using laboratory experiments

Since OSIRIS started acquiring high-resolution observations of the surface of the nucleus of comet 67P/Churyumov-Gerasimenko, over one hundred meter-sized bright spots have been identified in numerous types of geomorphologic regions, but mostly located in areas receiving low insolation. The bright spots are either clustered, in debris fields close to decameter-high cliffs, or isolated without structural relation to the surrounding terrain. They can be up to ten times brighter than the average surface of the comet at visible wavelengths and display a significantly bluer spectrum. They do not exhibit significant changes over a period of a few weeks. All these observations are consistent with exposure of water ice at the surface of boulders produced by dislocation of the weakly consolidated layers that cover large areas of the nucleus. Laboratory experiments show that under simulated comet surface conditions, analog samples acquire a vertical stratification with an uppermost porous mantle of refractory dust overlaying a layer of hard ice formed by recondensation or sintering under the insulating dust mantle. The evolution of the visible spectrophotometric properties of samples during sublimation is consistent with the contrasts of brightness and color seen at the surface of the nucleus. Clustered bright spots are formed by the collapse of overhangs that is triggered by mass wasting of deeper layers. Isolated spots might be the result of the emission of boulders at low velocity that are redepositioned in other regions.

Toxicity of cadmium to soil microbial activity: effect of sewage sludge addition to soil on the ecological dose

Cadmium has a toxic effect on soil microbial activity which plays an important role in nutrient cycling and, therefore, in maintaining soil fertility. In addition, the mobility of this heavy metal in soil is affected by the addition of urban wastes such as sewage sludge. This study was conducted to determine the effect of sewage sludge amendment of a semiarid soil, previously polluted with Cd, on the toxic effect of this heavy metal on soil microbial biomass and its activity. Dehydrogenase activity, ATP content, microbial soil respiration and microbial biomass carbon were used as bioindicators of the toxic effect of Cd. The inhibition of microbial activity and biomass by different Cd concentrations ranging from 0 to 8000 mg Cd kg −1 soil was described by three mathematical models in order to calculate three ecological doses of Cd: ED50 ,E D 10, and ED5 .I n general, higher ED values were calculated for the sewage sludge amended soil than for unamended soil. Thus the Cd toxicity to microbial activity of the sewage sludge amended soil can be considered lower than that of the unamended soil. Moreover, increased ED values with time after soil Cd contamination were observed.

Effect of composting on sewage sludges contaminated with heavy metals

The influence of composting on sewage sludges contaminated by Cd or by high concentrations of Cd, Cu, Ni and Zn was studied. Independently of the contamination which existed, the composting process led to a loss of weight, as shown by the decrease in organic matter; however, metal contamination did influence microbial populations and led to lower mineralization and humification being achieved. Contamination by Cd led to a much lower degree of nitrification during composting. Hydrolytic enzyme activities showed that large quantities of heavy metals had a negative effect on urease synthesis. Similarly, contamination negatively affected phosphatase; although not β-glucosidase, which increased with the level of contamination. The composting process rendered insoluble that portion of the metals which could be extracted with DTPA, CaCl2 or H2O by forming metal-humus complexes. Of the metals which were present at high quantity in the sewage sludges (Ni, Cu, Zn and Cd), Ni was the most easily extracted by H2O or CaCl2 and was therefore the most weakly bound to the organic matrix; Zn, in contrast, remained strongly bound.