Biodegradation and fate of ethyl tert-butyl ether (ETBE) in soil and groundwater: A review.

This review summarises the current state of knowledge on the biodegradation and fate of the gasoline ether oxygenate ethyl tert-butyl ether (ETBE) in soil and groundwater. Microorganisms have been identified in soil and groundwater with the ability to degrade ETBE aerobically as a carbon and energy source, or via cometabolism using alkanes as growth substrates. Aerobic biodegradation of ETBE initially occurs via hydroxylation of the ethoxy carbon by a monooxygenase enzyme, with subsequent formation of intermediates which include acetaldehyde, tert-butyl acetate (TBAc), tert-butyl alcohol (TBA), 2-hydroxy-2-methyl-1-propanol (MHP) and 2-hydroxyisobutyric acid (2-HIBA). Slow cell growth and low biomass yields on ETBE are believed to result from the ether structure and slow degradation kinetics, with potential limitations on ETBE metabolism. Genes known to facilitate transformation of ETBE include ethB (within the ethRABCD cluster), encoding a cytochrome P450 monooxygenase, and alkB-encoding alkane hydroxylases. Other genes have been identified in microorganisms but their activity and specificity towards ETBE remains poorly characterised. Microorganisms and pathways supporting anaerobic biodegradation of ETBE have not been identified, although this potential has been demonstrated in limited field and laboratory studies. The presence of co-contaminants (other ether oxygenates, hydrocarbons and organic compounds) in soil and groundwater may limit aerobic biodegradation of ETBE by preferential metabolism and consumption of available dissolved oxygen or enhance ETBE biodegradation through cometabolism. Both ETBE-degrading microorganisms and alkane-oxidising bacteria have been characterised, with potential for use in bioaugmentation and biostimulation of ETBE degradation in groundwater.

Influence of contaminant exposure on the development of aerobic ETBE biodegradation potential in microbial communities from a gasoline-impacted aquifer.

Aerobic biodegradation of ethyl tert butyl ether (ETBE) in a gasoline-impacted aquifer was investigated in laboratory microcosms containing groundwater and aquifer material from ETBE-impacted and non-impacted locations amended with either ETBE, or ETBE plus methyl tert butyl ether (MTBE). As sole substrate, ETBE was biodegraded (maximum rate of 0.54 day-1) without a lag in ETBE-impacted microcosms but with a lag of up to 66 days in non-impacted microcosms (maximum rate of 0.38 day-1). As co-substrate, ETBE was biodegraded preferentially (maximum rate of 0.25 and 0.99 day-1 in non-impacted and impacted microcosms, respectively) before MTBE (maximum rate of 0.24 and 0.36 day-1 in non-impacted and impacted microcosms, respectively). Further addition of ETBE and MTBE reduced lags and increased biodegradation rates. ethB gene copy numbers increased significantly (>100 fold) after exposure to ETBE, while overall cell numbers remained constant, suggesting that ethB-containing microorganisms come to dominate the microbial communities. Deep sequencing of 16S rRNA genes identified members of the Comamonadaceae family that increased in relative abundance upon exposure to ETBE. This study demonstrates the potential for ETBE biodegradation within the unsaturated and saturated zone, and that ETBE biodegrading capability is rapidly developed and maintained within the aquifer microbial community over extended timescales.

Influence of artificial drainage system design on the nitrogen attenuation potential of gley soils: Evidence from hydrochemical and isotope studies under field-scale conditions.

In North Atlantic Europe intensive dairy farms have a low nitrogen (N) use efficiency, with high N surpluses often negatively affecting water quality. Low feed input systems on heavy textured soils often need artificial drainage to utilise low cost grassland and remain profitable. Heavy textured soils have high but variable N attenuation potential, due to soil heterogeneity. Furthermore, drainage system design can influence the potential for N attenuation and subsequent N loadings in waters receiving drainage from such soils. The present study utilises end of pipe, open ditch and shallow groundwater sampling points across five sites in SW Ireland to compare and rank sites based on N surplus, water quality and "net denitrification", and to develop a conceptual framework for the improved management of heavy textured dairy sites to inform water quality N sustainability. This includes both drainage design and "net denitrification" criterion, as developed within this study.N surplus ranged from 211 to 292 kg N/ha (mean of 252 kg N/sourha) with a common source of organic N across all locations. The predicted soil organic matter (SOM) N release potential from top-subsoil layers was high, ranging from 115 to >146 kg N/ha. Stable isotopes analyses showed spatial variation in the extent of specific N-biotransformation processes, according to drainage location and design. Across all sites, nitrate (NO3-N) was converted to ammonium (NH4+-N), which migrated offsite through open ditch and shallow groundwater pathways. Using the ensemble data the potential for soil N attenuation could be discriminated by 3 distinct groups reflecting the relative dominance of in situ N-biotransformation processes deduced from water composition: Group 1 (2 farms, ranked with high sustainability, NH4+ < 0.23 mg N/l, δ15N-NO3- > 5‰ and δ18O-NO3- > 10‰), low NH4+-N concentration coupled with a high denitrification potential; Group 2 (1 farm with moderate sustainability, NH4+ < 0.23 mg N/l, δ15N-NO3- < 8‰ and δ18O-NO3- < 8‰), low NH4+-N concentration with a high nitrification potential and a small component of complete denitrification; Group 3 (2 farms, ranked with low sustainability, NH4+ > 0.23 mg N/l, 14‰ > Î´15N-NO3- > 5‰ and 25‰ > Î´18O-NO3- > -2‰), high NH4+-N concentration due to low denitrification. The installation of a shallow drainage system (e.g. mole or gravel moles at 0.4 m depth) reduced the "net denitrification" ranking of a site, leading to water quality issues. From this detailed work an N sustainability tool for any site, which presents the relationship between drainage class, drainage design (if present), completeness of denitrification, rate of denitrification and NH4-N attenuation was developed. This tool allows a comparison or ranking of sites in terms of their N sustainability. The tool can also be used pre-land drainage and presents the consequences of future artificial land drainage on water quality and gaseous emissions at a given site.

Infection of Arabidopsis thaliana leaves with Albugo candida (white blister rust) causes a reprogramming of host metabolism.

Abstract Albugo candida (Pers.) (O.) Kunze is a biotrophic pathogen which infects the crucifer Arabidopsis thaliana (L.) Heynh forming discrete areas of infection. Eight days after inoculation of leaves, white blisters became visible on the under surface of the leaf although no symptoms were apparent on the upper surface. By day 14, the region of leaf invaded by fungal mycelium had become chlorotic. Recently it has been hypothesized that an accumulation of soluble carbohydrates, following an increase in invertase activity, may trigger sugar signal transduction pathways leading to the repression of photosynthetic gene expression and to the induction of defence proteins. This hypothesis was investigated by quantifying localized changes in carbohydrate and photosynthetic metabolism and the expression of genes encoding photosynthetic and defence proteins. Quantitative imaging of chlorophyll fluorescence revealed that the rate of photosynthesis declined progressively in the invaded regions of the leaf. However, in uninfected regions of the infected leaf the rate of photosynthesis was similar to that measured in the control leaf until late on during the infection cycle when it declined. Images of nonphotochemical fluorescence quenching (NPQ) suggested that the capacity of the Calvin cycle had been reduced in infected regions and that there was a complex metabolic heterogeneity within the infected leaf. A. candida also caused localized changes in the carbohydrate metabolism of the leaf; soluble carbohydrates accumulated in the infected region whereas the amount of starch declined. The reverse was seen in uninfected regions of the infected leaf; carbohydrates did not accumulate until late on during infection and the amount of starch increased as the infection progressed. There was an increase in the activity of invertases which was confined to regions of the leaf invaded by the fungal mycelium. The increase in apoplastic invertase activity was of host origin, as mRNA levels of the ATbetaFRUCT1 gene (measured by semiquantitative RT-PCR) increased 40-fold in the infected region. The increase in soluble invertase activity resulted from the appearance of a new isoform in the invaded region of the leaf. Current evidence suggests that this was of fungal origin. Northern blot analysis of cab and rbcS showed that photosynthetic gene expression was repressed in the infected leaf from 6 days after inoculation (DAI) when compared to control leaves. In contrast, there was no detectable induction of defence proteins in the infected leaf. These data are discussed in the context of the sugar-sensing hypothesis presented above.

Purification and characterisation of soluble invertases from leaves of Arabidopsis thaliana.

Multiple isoforms of beta-fructofuranosidase (invertase, EC 3.2.1.26) were identified in mature green leaves of the cruciferous plant Arabidopsis thaliana (L.) Heynh. There were four major and one minor isoforms of soluble acid invertase and an additional activity which could be released from the cell wall by buffers of high ionic strength. This study reports the separation and characterisation of three soluble isoforms following ammonium sulphate and polyethylene glycol 6000 precipitations, Concanavalin A, MonoQ ion exchange, Superose 12 size-exclusion chromatography and chromatofocusing. These isoforms, designated INV1, INV2 and INV3, had isoelectric points of 4.75, 4.70 and 4.65 and a Km for sucrose of 5, 12 and 5 mM, respectively. Each had a pH optimum of 5.5, exhibited optimal activity at 45 degrees C and used sucrose as the preferred substrate. All fractions containing these isoforms contained a 52-kDa polypeptide which was specifically detected by immunoblotting with an antibody raised against deglycosylated wheat invertase. The N-terminal amino-acid sequence of this polypeptide was homologous to acid invertases isolated from other plant species. The possible origin of isoforms of soluble acid invertase is discussed.

Deletion analysis of a phytochrome-regulated monocot rbcS promoter in a transient assay system.

We have developed a transient gene expression assay system in the aquatic monocot Lemna gibba in which DNA was introduced into intact tissue by particle bombardment. Constructs based on the Lemna rbcS gene SSU5B, which is positively regulated by phytochrome in vivo, also showed phytochrome regulation in the transient assay system. Reporter gene expression increased 12-fold over dark levels in response to a single treatment with red light. This increase was not observed if far-red light was immediately followed by the red light. A 5' deletion analysis of the promoter defined a region from position -205 to position -83 relative to the start of transcription as necessary to observe the phytochrome response. This region contains the binding site for the light-induced binding activity (LRF-1) found in Lemna nuclear extracts. Upstream of position -205, we found evidence for the presence of at least two upstream activating sequences and a silencer.

Differential expression of individual genes encoding the small subunit of ribulose-1,5-bisphosphate carboxylase in Lemna gibba.

The gene family encoding the small subunit (SSU) of ribulose-1,5-bisphosphate carboxylase/oxygenase in the monocot Lemna gibba contains approximately twelve members. We have isolated six of these genes from a genomic library, and sequenced five of the coding regions. The transit peptide nucleotide sequences are conserved, but less highly than the mature polypeptide coding sequence. The mature polypeptide amino acid sequences are identical to each other and to the sequence deduced from a cDNA clone derived from a seventh gene. Each of the five fully characterized genomic sequences contains a single intron in precisely the same position as the second intron of several dicots. The intron sequences differ in length and are less conserved than the coding sequences. The 3'-untranslated regions of the different genes have been sequenced and used to prepare gene-specific probes. These probes have been used to study the expression levels of individual rbcS sequences. Expression of six of the seven genes can be detected in total RNA isolated from plants grown in continuous light. The levels of RNA encoded by each expressed gene are regulated by the action of phytochrome, but there is variability in the amount of expression of each RNA.

Effects of the similarity and dissimilarity between familiarization and test objects on recognition memory in infants following unimodal and bimodal familiarization.

Recognition memory for the shape of an object following unimodal (visual) and bimodal (visual and haptic) familiarization was investigated in 2 experiments. In both experiments, 6-month-old infants were familiarized either unimodally or bimodally with simple objects. In the first experiment the subjects were tested unimodally after familiarization, and in the second the subjects were tested bimodally. Visual recognition memory was only evident when the conditions of familiarization and testing were identical. 2 possible explanations of the data are presented, and the implications of these for the conclusions generated by previous studies are discussed.