Survival of Escherichia coli in stormwater biofilters.

Biofilters are widely adopted in Australia for stormwater treatment, but the reported removal of common faecal indicators (such as Escherichia coli (E. coli)) varies from net removal to net leaching. Currently, the underlying mechanisms that govern the faecal microbial removal in the biofilters are poorly understood. Therefore, it is important to study retention and subsequent survival of faecal microorganisms in the biofilters under different biofilter designs and operational characteristics. The current study investigates how E. coli survival is influenced by temperature, moisture content, sunlight exposure and presence of other microorganisms in filter media and top surface sediment. Soil samples were taken from two different biofilters to investigate E. coli survival under controlled laboratory conditions. Results revealed that the presence of other microorganisms and temperature are vital stressors which govern the survival of E. coli captured either in the top surface sediment or filter media, while sunlight exposure and moisture content are important for the survival of E. coli captured in the top surface sediment compared to that of the filter media. Moreover, increased survival was found in the filter media compared to the top sediment, and sand filter media was found be more hostile than loamy sand filter media towards E. coli survival. Results also suggest that the contribution from the tested environmental stressors on E. coli survival in biofilters will be greatly affected by the seasonality and may vary from one site to another.

Evaluating Escherichia coli removal performance in stormwater biofilters: a preliminary modelling approach.

Stormwater biofilters are not currently optimised for pathogen removal since the behaviour of these pollutants within the stormwater biofilters is poorly understood. Modelling is a common way of optimising these systems, which also provides a better understanding of the major processes that govern the pathogen removal. This paper provides an overview of a laboratory-scale study that investigated how different design and operational conditions impact pathogen removal in the stormwater biofilters. These data were then used to develop a modelling tool that can be used to optimise the design and operation of the stormwater biofilters. The model uses continuous simulations where adsorption and desorption were dominant during wet weather periods and first order die-off kinetics were significant in dry periods between the wet weather events. Relatively high Nash Sutcliffe Efficiencies (>0.5) indicate that the calibrated model is in good agreement with observed data and the optimised model parameters were comparable with values reported in the literature. The model's sensitivity is highest towards the adsorption process parameter followed by the die-off and desorption rate parameters, which implies that adsorption is the governing process of the model. Vegetation is found to have an impact on the wet weather processes since the adsorption and desorption parameters vary significantly with the different plant configurations. The model is yet to be tested against field data and needs to be improved to represent the effect of some other biofilter design configurations, such as the inclusion of the submerged zone.

Evaluating Escherichia coli removal performance in stormwater biofilters: a laboratory-scale study.

Biofilters are common, low energy technologies used for the treatment of urban stormwater. While they have shown promising results for the removal of stormwater microorganisms, certain factors affect their performance. Hence, this study investigated the effects of particle-microbial interaction, inflow concentration, antecedent microbial levels and plant species on microbial removal capacity. A biofilter column study was set up to evaluate removal performance and a sequential filtration procedure was used to estimate microbial partitioning. The columns were dosed with different concentrations of free phase Escherichia coli only and E. coli mixed with stormwater sediment. Results indicate that the microbial removal is significantly affected by inflow concentration and antecedent microbial levels. Leaching was only observed when a relatively low inflow concentration event occurred within a short period after a very high inflow concentration event. Finally, Lomandra longifolia showed better removal compared with Carex appressa.

Pharmacokinetics of muraglitazar (BMS-298585), a dual peroxisome proliferator-activated receptors (PPAR) alpha and gamma activator, in mice, rats, dogs, and monkeys.

The pharmacokinetic parameters of muraglitazar, a novel dual-activator of the peroxisome proliferator-activated receptors (PPAR) alpha and gamma, were determined in mice, rats, dogs, and monkeys after intravenous and oral administration. In the mouse, rat, and monkey the absolute oral bioavailability of muraglitazar ranged from 64 to 88%, and in the dog oral bioavailability was approximately 18%. The systemic clearance values of muraglitazar in the mouse, rat, dog, and cynomolgus monkey were 1.2, 3.0, 12.3 and 1.2 ml min-1 kg-1, respectively. The terminal elimination half-life was 2.4 h in dogs and 7.3 h in rats. The terminal elimination half-life could not be determined in the mouse and monkey because the sampling interval did not adequately cover the terminal elimination phase. Muraglitazar appears to be distributed outside of the vasculature, with the steady-state volume of distribution being approximately twofold that of the vascular volume in rats and dogs, and approximately twofold that of the total body water in mice. The systemic plasma clearance of muraglitazar in humans was predicted to be approximately 12-14 ml min-1 kg-1 based on allometry or by scaling of in vitro clearance parameters. Overall, the pharmacokinetic parameters of muraglitazar in preclinical species were acceptable for the advancement of the compound as a clinical candidate.

Functional expression of human intestinal Na+-dependent and Na+-independent nucleoside transporters in Xenopus laevis oocytes.

We have shown previously that the human jejunal brush border membrane expresses both the N1 (cif) and the N2 (cit) Na+-dependent (concentrative) nucleoside transporters but not the Na+-independent (facilitative) nitrobenzylmercaptopurineriboside (NBMPR)-sensitive (es) transporter (Patil SD and Unadkat JD, Am J Physiol, 272: 1314-1320, 1997). In the present study, we have demonstrated that when Xenopus laevis oocytes are microinjected with human jejunal mRNA, four nucleoside transporters are expressed simultaneously, namely the N1 and N2 Na+-dependent nucleoside transporters and the es and the NBMPR-insensitive (ei) Na+-independent transporters. The expressed Na+-dependent nucleoside transporters showed substrate specificity identical to that previously described by us using jejunal brush border membrane vesicles (Patil SD and Unadkat JD, Am J Physiol, 272: 1314-1320, 1997). The expressed es and ei Na+-independent transporters demonstrated broad substrate selectivity with both purines and pyrimidines capable of inhibiting the uptake of guanosine and thymidine mediated by this transporter. The expressed Na+-dependent nucleoside transporters mediated the transport of their respective nucleoside substrates with a high affinity and a low capacity, whereas the es and the ei transporters mediated the transport of nucleosides with a low affinity and a high capacity. Collectively, these observations suggest that the Na+-independent nucleoside transporters are expressed in the basolateral membrane of the human jejunal epithelium. Based on these data, we hypothesize that the concentrative transporters in the brush border membrane and equilibrative transporters in the basolateral membrane are arranged in series in the human jejunal epithelium to allow efficient vectorial transport of nucleosides from the lumen to the blood. The simultaneous expression of four nucleoside transporters in X. laevis oocytes establishes a basis for molecular cloning of these four human nucleoside transporters.

Cloning and sequencing of a full-length rat sucrase-isomaltase-encoding cDNA.

Sucrase-isomaltase (SI) has been widely used as a marker enzyme to study cellular differentiation in the small intestine. We isolated a 6.1-kb SI cDNA clone (GC1.4) from a size-fractionated cDNA library from rat intestine. Sequencing of this cDNA clone showed 6066 nucleotides (nt) with an open reading frame (ORF) of 1841 amino acids (aa). The nt sequence correctly predicts several known aa stretches in the protein. The deduced as sequence showed 78 and 75% overall identity with the rabbit and human SI, respectively. At the active sites of both S and I, the rat nt sequence encodes stretches of 14 and 16 aa, respectively, which show 100% identity to rabbit and human SI. In the region immediately beyond the transmembrane domain, the rat sequence encodes an extra 10 aa, as compared to rabbit and human. This 10-aa insertion consists almost entirely of Pro, Ser and Thr, and may be responsible for additional O-glycosylations of rat SI. The cDNA contains a 3'-UTR (untranslated region) of 499 nt with polyadenylation signal sequence and a poly(A) tract. The ATG start codon was found 41 nt downstream from the 5' end of the cDNA. Primer extension experiments showed the cap site to be 61 nt upstream from the start codon. The results indicate that our cDNA clone lacks only 20 nt in the 5'-UTR. Given that this cDNA encodes the entire coding region of SI, it should be useful in elucidating the regulatory mechanisms of SI biosynthesis, localization and targeting during rat intestinal development and differentiation.

Comparison of rates of intracellular metabolism of zidovudine in human and primate peripheral blood mononuclear cells.

3'-Azido-3'-deoxythymidine (AZT) is a drug of choice for the treatment of AIDS. On the basis of pharmacokinetic data, the nonhuman primate Macaca nemestrina has been shown to be a suitable animal model for use in the study of the disposition of AZT. However, since AZT is activated to its metabolite, the AZT triphosphate (AZTTP), intracellularly, we investigated the intracellular activation of AZT in peripheral blood mononuclear cells (PBMCs) of healthy and simian immunodeficiency virus-infected macaques and compared it with that in PBMCs obtained from human volunteers. At 5 microM extracellular AZT, both human and macaque PBMCs rapidly convert AZT to AZT monophosphate (AZTMP) (84% of total phosphorylated products) in 4 h. Increases in AZTMP levels of 7.7- and 12-fold were observed in human and macaque PBMCs, respectively, when the extracellular AZT concentration increased from 0.45 to 14.4 microM. Similar ratios of AZT metabolites, AZT diphosphate (AZTDP)/AZTTP (0.7 to 1.4), AZTMP/AZTDP (3 to 14), and AZTMP/AZTTP (3 to 19), over the same AZT concentration range were observed in both human and macaque PBMCs, suggesting that these cells have similar capacities to phosphorylate AZT. Simian immunodeficiency virus-infected macaque PBMCs showed a fivefold increase in intracellular AZT and AZTMP levels and a twofold increase in AZTDP and AZTTP levels (picomoles per 10(7) cells) when compared with those in the uninfected cells (at 4 h with 0.9 microM extracellular concentration). This increase in AZT metabolite levels has also been reported for human immunodeficiency virus-infected PBMCs. Collectively, given the similarities in phosphorylation profiles between healthy and infected human and macaque PBMCs, we conclude that the macaque is a suitable animal model for use in the study of factors that can effect the in vivo phosphorylation of AZT.

Expression of sucrase-isomaltase mRNA along the villus-crypt axis in the rat small intestine.

Sucrase-isomaltase has been used as a marker enzyme to study cell differentiation along the intestinal villus-crypt axis. Previous studies are in agreement that sucrase activity is confined to villus epithelial cells. However, immunoreactivity data are at conflict, with some studies reporting sucrase antigen in crypts as well as villi. To resolve this discrepancy, our goal was to determine the distribution of sucrase-isomaltase mRNA. A cDNA clone representing 3.0 kb of rat sucrase-isomaltase, including the sucrase active site, was characterized. Northern analysis of 12 tissues demonstrated a 6 kb transcript only in the small intestine. Jejunal cell fractions prepared by a washing technique showed declining levels of both sucrase activity and sucrase-isomaltase mRNA as well as increasing levels of thymidine kinase activity from early to later fractions. Since later fractions did not yield pure crypt cells, in situ hybridization using an 35S-labeled sucrase-isomaltase riboprobe was performed. The transition from zero to intense signal at the crypt-villus junction leads us to conclude that in the adult rat, sucrase-isomaltase gene expression is initiated only after cells leave the proliferative cycle and migrate onto the villi.

Radiation-induced recovery processes in cultured marsupial cells.

The ultraviolet sensitivity of Potorous tridactylus male kidney (PtK-2) cells is markedly increased by post irradiation treatment for 24 h with 5 microM emetine (which inhibits protein synthesis by acting on the 40S ribosomal subunit), or with 5 microM cycloheximide (which inhibits by interaction with the 60S subunit), or with the RNA polymerase II inhibitor 5,6-dichloro-1-beta-ribofuranosylbenzimidazole at 50 microM. All 3 treatments give the same sensitivity, while unirradiated cells are little affected. Shortening the time of treatment, or delaying application of the drugs decreases their effect on the same time schedule. Preiirradiation of cells, with no drug treatment in the following 8 h, diminishes the sensitivity to a subsequent irradiation with protein synthesis blocked afterwards. Photoreactivation immediately following such preirradiation eliminates its desensitizing effect. Inhibiting protein synthesis after irradiation also markedly reduces the frequency of UV-induced mutants in the surviving population. These facts suggest that gene expression in the period following irradiation facilitates recovery from radiation damage, with an increased probability of mutation, reminiscent of the "SOS response" in Escherichia coli.