Coliform concentration reduction and related performance evaluation of a down-flow anaerobic fixed bed reactor treating low-strength saline wastewater

Low-strength saline wastewater may be generated by tourist facilities, industries and communities located in coastal areas. Sea salts, mostly chlorides, when present in wastewaters at high concentrations, can cause inhibition on biological treatment processes. In this study, a laboratory down-flow anaerobic fixed bed reactor (DFAFBR) was used for treating saline wastewater. This wastewater was simulated by dilution of piggery manure in a synthetic saline water to obtain a final total COD concentration in the range of 1100-2900 mg/l and a salt concentration of 15 g/l. The DFAFBR was operated at hydraulic retention times (HRT) of 96, 48, 24 and 12 h. The results showed that at sea salts concentrations in the range from 5 to 15 g/l, total coliform concentration reduction efficiencies higher than 97% were achieved. A decrease in the total and faecal coliform concentration reduction efficiencies from 99.5% to 90.5% and 92.5%, respectively, was observed when the HRT decreased from 96 to 12 h. Enumeration of coliform bacteria isolated from the biofilm in different zones of the reactor showed that more than 94% of the total amount was removed in the upper zone. A HRT of 24 h was required to obtain total COD, organic-N, total-P and faecal coliform concentration reduction efficiencies higher than 72%, 51%, 39% and 98%, respectively. A concentration of 8.4 g/l for chlorides, 1.25 g/l for sulphates and 4.6 g/l for sodium did not affect the process performance.     

Construction and characterization of a minimized version of the HIV-1 pNL4-3 plasmid and its application for pseudotyping HIV-1 vectors

The pUC-based pNL4-3 plasmid is the most widely used vector for in vitro manipulations of the HIV-1 proviral sequences. We have developed a minimal plasmid (pCHUS) based on pNL4-3, which may be useful to facilitate the design of HIV-based constructions. The strategy that has allowed us to construct pCHUS includes the following steps: (1) pNL-3 digestion by using restriction sites contained within the long terminal repeats (LTRs), (2) recircularization of the fragment containing the pUC18 sequence, (3) amplification of the LTR region restored in the previous step, (4) double digestion of the products obtained in steps 2 and 3, (5) ligation of the fragment containing ColE1+Amp^R with the LTR fragment, (6) linearization of the intermediate plasmid obtained, and (7) insertion of the fragment containing the proviral genome into the linearized vector. The pCHUS plasmid includes essential information for its replication and antibiotic selection in bacteria, but it lacks all the unnecessary sequences. Our results suggest that pCHUS may be more advantageous than pNL4-3 for in vitro manipulation of the HIV-1 proviral genome. In addition, we describe a potential application of this new vector for pseudotyping HIV-1 particles, using a single plasmid transfection, as a more helpful alternative to the traditionally used cotransfection method.     

Structural and hemostatic activities of a sulfated galactofucan from the brown alga Spatoglossum schroederi. An ideal antithrombotic agent?

The brown alga Spatoglossum schroederi contains three fractions of sulfated polysaccharides. One of them was purified by acetone fractionation, ion exchange, and molecular sieving chromatography. It has a molecular size of 21.5 kDa and contains fucose, xylose, galactose, and sulfate in a molar ratio of 1.0:0.5:2.0:2.0 and contains trace amounts of glucuronic acid. Chemical analyses, methylation studies, and NMR spectroscopy showed that the polysaccharide has a unique structure, composed of a central core formed mainly by 4-linked beta-galactose units, partially sulfated at the 3-O position. Approximately 25% of these units contain branches of oligosaccharides (mostly tetrasaccharides) composed of 3-sulfated, 4-linked alpha-fucose and one or two nonsulfated, 4-linked beta-xylose units at the reducing and nonreducing end, respectively. This sulfated galactofucan showed no anticoagulant activity on several "in vitro" assays. Nevertheless, it had a potent antithrombotic activity on an animal model of experimental venous thrombosis. This effect is time-dependent, reaching the maximum 8 h after its administration compared with the more transient action of heparin. The effect was not observed with the desulfated molecule. Furthermore, the sulfated galactofucan was 2-fold more potent than heparin in stimulating the synthesis of an antithrombotic heparan sulfate by endothelial cells. Again, this action was also abolished by desulfation of the polysaccharide. Because this sulfated galactofucan has no anticoagulant activity but strongly stimulates the synthesis of heparan sulfate by endothelial cells, we suggested that this last effect may be related to the "in vivo" antithrombotic activity of this polysaccharide. In this case the highly sulfated heparan sulfate produced by the endothelial cells is in fact the antithrombotic agent. Our results suggested that this sulfated galactofucan may have a potential application as an antithrombotic drug.     

Cyanide-resistant respiration is frequent, but confined to yeasts incapable of aerobic fermentation

In Pichia membranifaciens, cyanide-resistant respiration (CRR) sensitive to salicylhydroxamic acid emerged after forced aeration of starved cells for 4 h. Surveying a large number of species by this simple methodology, we found that CRR is very frequent among yeasts. Remarkably, considering our results together with previous data in the literature, CRR was present in 24 out of 28 non-fermentative or Crabtree-negative yeasts and absent in 10 out of 12 Crabtree-positive yeasts. We submit that, as alternatives to cytochromic respiration, yeasts developed two strategies: either aerobic fermentation in Crabtree-positive yeasts or CRR in non-fermentative or Crabtree-negative yeasts.     

Damage induction by direct electric current in tumoural target cells

Damage induction to tumour target cells (P815) by direct electric current (DC) was investigated. A 6 min treatment of P815 cells with DC generated decreased levels of cell viability and proliferation. The ultrastructural analysis of DC-treated cells revealed the presence of blebs, loss of cell surface filopodia, and ruptures in cell membrane. Mitochondrial alterations, swelling of cells, cytoplasmic matrix rarefaction, and cellular debri formation were also observed. The study shows that tumoural target cells can be damaged by direct electric current and this approach may provide means to understand the mechanism of tumour regression induced by electrochemical therapy.     

Effect of brown spider venom on basement membrane structures

Loxoscelism or necrotic arachnidism are terms used to describe lesions and reactions induced by bites (envenomation) from spiders of the genus Loxosceles. Envenomation has been reported to provoke dermonecrosis and haemorrhage at the bite site and haemolysis, disseminated intravascular coagulation and renal failure. The purpose of this work was to study the effect of the venom of the brown spider Loxosceles intermedia on basement membrane structures and on its major constituent molecules. Light microscopy observations showed that L. intermedia venom obtained through electric shock, which reproduces two major signals of Loxoscelism in the laboratory, exhibits activity toward basement membrane structures in mouse Engelbreth-Holm-Swarm (EHS) sarcoma. Basement degradation was seen by a reduced periodic acid-Schiff (PAS) and alcian blue staining as well as by a reduced immunostaining for laminin when compared to control experiments. Electron microscopy studies confirmed the above results, showing the action of the venom on EHS-basement membranes and demonstrating that these tissue structures are susceptible to the venom. Using purified components of the basement membrane, we determined through SDS-PAGE and agarose gel that the venom is not active toward laminin or type IV collagen, but is capable of cleaving entactin and endothelial heparan sulphate proteoglycan. In addition, when EHS tissue was incubated with venom we detected a release of laminin into the supernatant, corroborating the occurrence of some basement membrane disruption. The venom-degrading effect on entactin was blocked by 1,10-phenanthroline, but not by other protease inhibitors such as PMSF, NEM or pepstatin-A. By using light microscopy associated with PAS staining we were able to identify that 1,10-phenanthroline also inhibits EHS-basement membrane disruption evoked by venom, corroborating that a metalloprotease of venom is involved in these effects. Degradation of these extracellular matrix molecules and the observed susceptibility of the basement membrane could lead to loss of vessel and glomerular integrity, resulting in haemorrhage and renal problems after envenomation.     

Assisted reproductive technology (ART) in humans: facts and uncertainties

Since the first in vitro fertilization (IVF) in human, the number of patients using Assisted Reproductive Technologies (ART) has increased tremendously. ART technologies have increased in number and their spectrum has also widened. The first IVF babies are now more than 20 years old. All the retrospective analyses have demonstrated that the obstetrical and pediatrical impact has not really affected single births. The main problems observed occur with multiple pregnancies, including high costs for the couples and for society. The decrease in the number of embryos transferred has improved the situation and moreover does not impair the final results. IntraCytoplasmic Sperm Injection (ICSI) is a more debatable and questionable technique with a real negative genetic impact. The main problem is chromosome abnormalities more specifically related to the sex chromosomes. The question of a systematic genetic work-up on the patients entering ICSI programs is discussed. No negative impact of cryopreservation has been demonstrated even though some controversy arises from time to time. Pre-implantation Genetic Diagnosis (PGD) is now a interesting tool for patients carrying genetic defects. Blastocyst biopsy now has a future role in reproductive medicine. Gender selection through sperm sorting is also now a reality. As with the other developing bio-technologies related to reproduction, there are ethical questions. The decisions concerning these technologies do not belong solely to scientists but are rather a matter for society to decide.     

The effects of selective and nonselective cyclooxygenase inhibitors on endothelin-1-induced fever in rats

It was previously shown that sustained fever can be induced in rats by central injection of endothelin-1 (ET-1). This peptide appears to participate in the mechanism(s) of LPS-induced fever, which is reduced by pretreatments with ET(B) receptor antagonists. In this study, we compared the effects of a nonselective cyclooxygenase (COX) inhibitor, indomethacin, with those of two selective COX-2 inhibitors, celecoxib and lumiracoxib, on ET-1-induced fever in rats. Fever induced in conscious animals by ET-1 (1 pmol icv) or LPS (5 mug/kg iv) was prevented by pretreatments with celecoxib (5 and 10 mg/kg) or lumiracoxib (5 mg/kg) given by oral gavage 1 h before stimuli. Lower doses of celecoxib had partial (2.5 mg/kg) or no effect (1 mg/kg). Indomethacin (2 mg/kg ip) partially inhibited fever induced by LPS but had no effect on ET-1-induced fever. The levels of PGE(2) and PGF(2alpha) in the cerebrospinal fluid (CSF) of pentobarbital sodium-anesthetized rats were significantly increased 3 h after the injection of LPS or ET-1. The latter increase was abolished by celecoxib at all tested doses and by indomethacin. In conclusion, selective COX-2 inhibitors were able to prevent ET-1-induced fever, indicating a role for COX-2 in this phenomenon. However, the fact that reduced CSF PG levels obtained with indomethacin and a low dose of celecoxib are not accompanied by changes in fever induced by ET-1, along with the lack of inhibitory effects of indomethacin on ET-1 fever, suggests that the latter might also involve COX-2-independent mechanisms.     

Simultaneous nitrification and formaldehyde biodegradation in an activated sludge unit

The simultaneous removal of formaldehyde and ammonium in a lab-scale activated sludge unit was investigated. The unit was operated at a hydraulic retention time of 2.4 days with an ammonium concentration in the influent of 350 mg NH4+-N/L, maintaining the ammonium loading rate at 0.15 g NH4+-N/Ld during the operation time. However, the applied organic loading rate was increased stepwise by increasing the formaldehyde concentration from 26 up to 3168 mg/L, corresponding to 0.01-1.40 g COD/Ld. High formaldehyde removal efficiencies, around 99.5% (+/-0.38), were maintained at all the formaldehyde concentrations. Ammonium removal was also very high during the operation period, around 99.9% (+/-0.01). The ammonium concentration in the effluent was lower than 0.1 mg NH4+-N/L at all applied organic loading rates, indicating that there was no inhibition of nitrification by formaldehyde.