Identification and characterization of a HEPN-MNT family type II toxin–antitoxin in Shewanella oneidensis

Toxin–antitoxin (TA) systems are prevalent in bacteria and archaea. However, related studies in the ecologically and bioelectrochemically important strain Shewanella oneidensis are limited. Here, we show that SO_3166, a member of the higher eukaryotes and prokaryotes nucleotide-binding (HEPN) superfamily, strongly inhibited cell growth in S. oneidensis and Escherichia coli. SO_3165, a putative minimal nucleotidyltransferase (MNT), neutralized the toxicity of SO_3166. Gene SO_3165 lies upstream of SO_3166, and they are co-transcribed. Moreover, the SO_3165 and SO_3166 proteins interact with each other directly in vivo, and antitoxin SO_3165 bound to the promoter of the TA operon and repressed its activity. Finally, the conserved Rx4-6H domain in HEPN family was identified in SO_3166. Mutating either the R or H abolished SO_3166 toxicity, confirming that Rx4-6H domain is critical for SO_3166 activity. Taken together, these results demonstrate that SO_3166 and SO_3165 in S. oneidensis form a typical type II TA pair. This TA pair plays a critical role in regulating bacterial functions because its disruption led to impaired cell motility in S. oneidensis. Thus, we demonstrated for the first time that HEPN-MNT can function as a TA system, thereby providing important insights into the understanding of the function and regulation of HEPNs and MNTs in prokaryotes.     

The effect of cyclic anaerobic–aerobic conditions on biodegradation of azo dyes

The effect of cyclic anaerobic–aerobic conditions on the biodegradative capability of the mixed microbial culture for the azo dye Remazol Brilliant Violet 5R (RBV-5R) was investigated in the sequencing batch reactor (SBR) fed with a synthetic textile wastewater. The SBR had a 12-h cycle time with anaerobic–aerobic periods of 3/9, 6/6 and 9/3 h. General SBR performance was assessed by measurement of catabolic enzymes (catechol 2,3-dioxygenase, azo reductase), chemical oxygen demand (COD), color and amount of aromatic amines. In this study, under steady-state conditions, the anaerobic period of the cyclic SBR was found to allow the reductive decolorization of azo dye. Longer anaerobic periods resulted in higher color removal efficiencies, approximately 71% for the 3-h, 87% for 6-h and 92% for the 9-h duration. Total COD removal efficiencies were over 84% under each of the cyclic conditions and increased as the length of the anaerobic period was increased; however, the highest color removal rate was attained for the cycle with the shortest anaerobic period of 3 h. During the decolorization of RBV-5R, two sulfonated aromatic amines (benzene based and naphthalene based) were formed. Additionally, anaerobic azo reductase enzyme was found to be positively affected with the increasing duration of the anaerobic period; however; it was vice versa for the aerobic catechol 2,3-dioxygenase (C23DO) enzyme.     

Effect of influent pH and alkalinity on the removal of chlorophenols in sequential anaerobic–aerobic reactors

This study was carried out to determine the effect of influent pH and alkalinity on the performance of sequential UASB and RBC reactors for the removal of 2-CP and 2,4-DCP from two different simulated wastewaters. The performance of methanogens at low (<6.0) to high (>8.0) pH values and at sufficiently high alkalinity (1500–3500 mg/l as CaCO₃) is described in this paper. Sequential reactors were capable of handling wastewaters with influent pH, 5.5–8.5. However, with influent pH 7.0 ± 0.1 UASB reactor showed best performance for 2-CP (99%) and 2,4-DCP (88%) removals. Increase in alkalinity/COD ratio in the influent (>1.1) caused gradual decrease in the chlorophenol removal in UASB reactors. The UASB reactors could not tolerate wastewater with higher alkalinity/COD ratio (2.6) and showed significant deterioration of its performance in terms of chlorophenols removal achieving only 74.7% 2-CP and 60% 2,4-DCP removals, respectively.     

Sequential anaerobic–aerobic degradation of munitions waste

A sequential anaerobic–aerobic biodegradation of hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX) was studied. The results demonstrated that: (i) a complete degradation of RDX was achieved within 20 days using a consortium of bacteria from a wastewater activated sludge, (ii) RDX degradation did not occur under aerobic conditions alone, (iii) RDX-degrading bacterial strain that was isolated from the activated sludge completely degraded RDX within 2 days, and (iv) RDX- induced protein expressions were observed in the RDX-degrading bacterial strain. Based on fatty acid composition and a confirmation with a 16S rRNA analysis, the RDX-degrading bacterial strain was identified as a Bacillus pumilus—GC subgroup B.     

The effect of biological sulfate reduction on anaerobic color removal in anaerobic–aerobic sequencing batch reactors

Combination of anaerobic–aerobic sequencing processes result in both anaerobic color removal and aerobic aromatic amine removal during the treatment of dye-containing wastewaters. The aim of the present study was to gain more insight into the competitive biochemical reactions between sulfate and azo dye in the presence of glucose as electron donor source. For this aim, anaerobic–aerobic sequencing batch reactor fed with a simulated textile effluent including Remazol Brilliant Violet 5R (RBV 5R) azo dye was operated with a total cycle time of 12 h including anaerobic (6 h) and aerobic cycles (6 h). Microorganism grown under anaerobic phase of the reactor was exposed to different amounts of competitive electron acceptor (sulfate). Performance of the anaerobic phase was determined by monitoring color removal efficiency, oxidation reduction potential, color removal rate, chemical oxygen demand (COD), color, specific anaerobic enzyme (azo reductase) and aerobic enzyme (catechol 1,2-dioxygenase), and formation of aromatic amines. The presence of sulfate was not found to significantly affect dye decolorization. Sulfate and azo dye reductions took place simultaneously in all operational conditions and increase in the sulfate concentration generally stimulated the reduction of RBV 5R. However, sulfate accumulation under anaerobic conditions was observed proportional to increasing sulfate concentration.     

Stability of cytochromes c′ from psychrophilic and piezophilic Shewanella species: implications for complex multiple adaptation to low temperature and high hydrostatic pressure

Extremophiles - The stability of dimeric cytochrome c′ from a thermophile, as compared with that of a homologous mesophilic counterpart, is attributed to strengthened interactions around the...     

Iron Triggers λSo Prophage Induction and Release of Extracellular DNA in Shewanella oneidensis MR-1 Biofilms

Prophages are ubiquitous elements within bacterial chromosomes and affect host physiology and ecology in multiple ways. We have previously demonstrated that phage-induced lysis is required for extracellular DNA (eDNA) release and normal biofilm formation in Shewanella oneidensis MR-1. Here, we investigated the regulatory mechanisms of prophage λSo spatiotemporal induction in biofilms. To this end, we used a functional fluorescence fusion to monitor λSo activation in various mutant backgrounds and in response to different physiological conditions. λSo induction occurred mainly in a subpopulation of filamentous cells in a strictly RecA-dependent manner, implicating oxidative stress-induced DNA damage as the major trigger. Accordingly, mutants affected in the oxidative stress response (ΔoxyR) or iron homeostasis (Δfur) displayed drastically increased levels of phage induction and abnormal biofilm formation, while planktonic cells were not or only marginally affected. To further investigate the role of oxidative stress, we performed a mutant screen and identified two independent amino acid substitutions in OxyR (T104N and L197P) that suppress induction of λSo by hydrogen peroxide (H₂O₂). However, λSo induction was not suppressed in biofilms formed by both mutants, suggesting a minor role of intracellular H₂O₂ in this process. In contrast, addition of iron to biofilms strongly enhanced λSo induction and eDNA release, while both processes were significantly suppressed at low iron levels, strongly indicating that iron is the limiting factor. We conclude that uptake of iron during biofilm formation triggers λSo-mediated lysis of a subpopulation of cells, likely by an increase in iron-mediated DNA damage sensed by RecA.     

Thermal stability of cytochrome c₅ of pressure-sensitive Shewanella livingstonensis

Cytochrome c? of pressure-sensitive Shewanella livingstonensis (SL cytc?) exhibits lower thermal stability than a highly homologous counterpart of pressure-tolerant Shewanella violacea. This stability difference is due to an enthalpic effect that can be attributed to the amino acid residue at position 50 (Leu or Lys). These cytc? proteins are appropriate materials for understanding the protein stability mechanism.     

Cycling treatment of anaerobic and aerobic incubation increases the content of γ-aminobutyric acid in tea shoots

Summary. γ-Aminobutyric acid (GABA), a hypotensive compound, is formed from glutamic acid under anaerobic condition in tea shoots. Glutamic acid was exhausted in the first three hours of anaerobic incubation and the increase of GABA stopped. After that, when tea shoots were released under aerobic condition, glutamic acid reproduced rapidly. After one hour of aerobic incubation, tea shoots were given three hours of anaerobic incubation again and then accumulated glutamic acid changed to GABA. The content of GABA increased much more than usual anaerobic incubation. GABA was more in the tea stem than in the leaf. Received January 4, 2000 Accepted March 1, 2000     

Biodegradation of azo dyes in a sequential anaerobic–aerobic system

A sequential anaerobic–aerobic treatment process based on mixed culture of bacteria isolated from textile dye effluent-contaminated soil was used to degrade sulfonated azo dyes Orange G (OG), Amido black 10B (AB), Direct red 4BS (DR) and Congo red (CR). Under anaerobic conditions in a fixed-bed column using glucose as co-substrate, the azo dyes were reduced and amines were released by the bacterial biomass. The amines were completely mineralized in a subsequent aerobic treatment using the same isolates. The maximum degradation rate observed in the treatment system for OG was 60.9 mg/l per day (16.99 mg/g glucose utilized), for AB 571.3 mg/l per day (14.46 mg/g glucose utilized), for DR 112.5 mg/l per day (32.02 mg/g glucose utilized) and for CR 134.9 mg/l per day (38.9 mg/g glucose utilized). Received: 6 August 1999 / Received revision: 20 December 1999 / Accepted: 24 December 1999     

Impacts of drought on leaf respiration in darkness and light in Eucalyptus saligna exposed to industrial-age atmospheric CO₂ and growth temperature

Our study assessed the impact of a wide range of industrial-age climate scenarios on leaf respiration (R) in Eucalyptus saligna. Well-watered or sustained drought-treated plants were grown in glasshouses differing in atmospheric CO? concentration ([CO?]) (280, 400 and 640 μl l?1) and temperature (26 and 30°C). Rates of R in darkness (R(dark) ) and light (R(light) ), photosynthesis (A) and related leaf traits (mass : area relationships, and nitrogen, phosphorus, starch and sugar concentrations) were measured. Light inhibited R in all cases (R(light) < R(dark) ) (well-watered: 40%; drought-treated: 73%). Growth [CO?] and temperature had little impact on area-based rates of R(dark) or R(light) , with R(light) exhibiting minimal thermal acclimation. By contrast, sustained drought resulted in reduced R(dark), R(light) and A, with the inhibitory effect of drought on A and R(light) (c. 50-70%) greater than that on R(dark) (c. 15%). Drought effects were fully reversible after watering. Variability in R(light) appeared to be dependent on the underlying rate of R(dark) and associated Rubisco activity. Collectively, our data suggest that there is an asynchronous response of leaf carbon metabolism to drought, and a tighter coupling between R(light) and A than between R(dark) and A, under both past and future climate scenarios. These findings have important implications for ecosystem/global models seeking to predict carbon cycling.     

The fate of lignin and lignin‐derived compounds in anaerobic environments

During ODP Leg 201 microbial communities in Eastern Equatorial Pacific Ocean and Peru Margin sediments were investigated. The sediment layers sampled extended down to 420 m below the sea floor, with estimated ages of up to 40 million years. Contamination-free anoxic slurries were inoculated into media containing different substrate combinations, all at micromolar concentration. These culture media were designed for a broad spectrum of physiological groups. A total of 162 pure cultures were isolated that could be grouped into 19 different phylotypes based on 16S rRNA gene analysis. The isolates belonged to the Alpha-, Gamma- and Deltaproteobacteria, the Firmicutes, Actinobacteria, and Bacteroidetes. The genera most frequently isolated were Bacillus (68 isolates) and Rhizobium (40 isolates). Comparison of strains with the same phylotypes by enterobacterial repetitive intergenic consensus (ERIC-PCR) analysis revealed the presence of several subgroups that did not correlate with medium, sediment depth or sampling site. The majority of the isolates, although obtained from anoxic environments and isolated under strictly anoxic conditions, turned out to be facultativly aerobic. Physiologically, the isolates were characterized as generalists, able to utilize a broad variety of electron donors with either oxygen, nitrate and in some cases manganese oxides as electron acceptors. The diversity inferred from physiological tests was even higher than that on the phylogenetic or genomic level. The outcome of the contamination tests, the isolation of close relatives of already known subsurface bacteria, the repeated finding of the same phylotype from different sites and the level of diversity present in the culture collection strongly suggest that indigenous deep-biosphere bacteria had been isolated.     

The effect of exogenous δ-aminolaevulinate on rat liver haem and cytochromes

The activity of delta-aminolaevulinate synthetase is generally regarded as rate-limiting for hepatic haem biosynthesis. It has been suggested that cytochrome synthesis may also be regulated by changes in delta-aminolaevulinate synthetase activity. This hypothesis was studied by injecting product, delta-aminolaevulinate, into adult rats over a 4-240h period. The concentrations of hepatic mitochondrial cytochromes a, b, c and c(1) were unchanged by treatment with delta-aminolaevulinate, allylisopropylacetamide or phenobarbital. In control animals, total microsomal haem content equalled the sum of cytochromes b(5) plus P-450. After delta-aminolaevulinate administration the total amount of microsomal haem, measured as the pyridine haemochromogen, exceeded these components, indicating the formation of a ;free' haem pool. Haem synthesis does not appear rate-limiting for hepatic cytochrome synthesis in the adult rat.     

Short‐term effect of hypoxia on ammonia excretion and respiration rates in the crab carcinus maenas

The metabolic response of the crab Carcinus maenas to short‐term hypoxia (60% and 35% saturated seawater) was studied at 17.5°C in fed, 3 day‐unfed and 6 day‐unfed crabs.

Ammonia excretion rate decreased under hypoxia: a 40% and 45% decrease in the normoxic rate was observed in fed crabs at 35% saturation and in 3 day‐unfed crabs at both hypoxic levels respectively. In the 6 day‐unfed crabs, the effect of hypoxia was concealed by the effect of starvation.

Oxygen consumption rate was directly related to the external O₂ tension irrespective of the crab's nutritional state. Stressed crabs behaved as a whole, as oxygen‐conformers.

A strong relationship was observed between ammonia excretion and oxygen consumption rates in fed crabs under hypoxia but not in starved crabs.     

Decolorization of anthraquinone dye intermediate and its accelerating effect on reduction of azo acid dyes by Sphingomonas xenophaga in anaerobic–aerobic process

Decolorization of 1-aminoanthraquinone-2-sulfonic acid (ASA-2) and its accelerating effect on the reduction of azo acid dyes by Sphingomonas xenophaga QYY were investigated. The study showed that ASA-2 could be efficiently decolorized by strain QYY under aerobic conditions according to the analysis of total organic carbon removal and UV-VIS spectra changes. Moreover, strain QYY was able to reduce azo acid dyes under anaerobic conditions. The effects of various operating conditions such as carbon sources, temperature, and pH on the reduction rate were studied. It was demonstrated that ASA-2 used as a redox mediator could accelerate the reduction process. Consequently the reduction of azo acid dyes mediated by ASA-2 and the decolorization of ASA-2 with strain QYY could be achieved in an anaerobic-aerobic process.     

Impacts of alien plants and man‐made disturbance on soil‐growing bryophyte and lichen diversity in coastal areas of Sardinia (Italy)

Abstract

Seventy phytosociological relevés were performed in 1 m × 1 m plots at 14 study sites spread along sandy shores in northern and southern Sardinia (Italy). The plots were selected in different habitat types (open dunes, native Juniperus woodlands, maquis, and plantations with Acacia, Eucalyptus and Pinus) according to a stratified sampling method in order to investigate impacts deriving from different levels of Carpobrotus spp. cover, dry litter from exotic trees, and other disturbance types. The quantile regression and logistic regression analyses revealed that the reduction in the amount of bryophyte and lichen cover on sand dunes of the study area is caused either by a high cover of Carpobrotus spp. mats or by a high cover of dry exotic litter in dense, unmanaged or poorly managed forest plantations. Additional detrimental effects are often driven by other kinds of man‐made disturbances. Forest management in the coastal areas of Sardinia should be gradually modified to take into account the conservation of bryophytes and lichens. Some of the biological indicators used are quite widespread in the Mediterranean coastal habitats or are exclusively associated with sand dunes; therefore, they can also be conveniently used as indicators of biological impacts in other countries or islands of the same biogeographical region.     

Biological sulphate reduction and redox mediator effects on azo dye decolourisation in anaerobic–aerobic sequencing batch reactors

In this work, the anaerobic period of an anaerobic–aerobic sequencing batch reactor was found to allow the reductive decolourisation of azo dyes. 1-l reactors were operated in 24-h cycles comprising anaerobic and aerobic reaction phases, fed with a simulated textile effluent including a reactive type (Remazol Brilliant Violet 5R) or an acid type (Acid Orange 7) azo dye. The aim was to assess the role of different redox phenomena in the anaerobic decolourisation process. Selective inhibition of sulphate reducing bacteria was carried out in the sulphate-containing, reactive dye fed reactor, resulting in nearly complete, though reversible and inhibition of decolourisation. The acid dye fed reactor's supplementation with sulphate, though resulting in sulphate reduction, did not improve decolourisation. Other redox mediators, namely quinones, were more effective in promoting electron transfer to the azo bond. Bio-augmentation of the acid dye fed reactor with a pure sulphate reducer strain known to decolourise azo dyes, Desulfovibrio alaskensis, was also carried out. Decolourisation was improved, but apparently as a result of the carbon source change required to support D. alaskensis growth. A chemically mediated reduction of the azo bond coupled to biological sulphate reduction, thus seemed to account for the high decolourisation yields of both dyes.     

Application of anaerobic–aerobic sequential treatment system to real textile wastewater for color and COD removal

A sequential anaerobic packed column reactor and an activated sludge unit was operated continuously for treatment of a textile industry wastewater, in Izmir, Turkey. Metal sponges were used as support material in anaerobic unit and pre-activated textile dyestuff biodegrading PDW facultative anaerobic bacterial culture was immobilized on the support particles. Effects of hydraulic retention times in anaerobic unit (θ_H anaerobic = 12–72 h) and initial COD concentration (COD₀ = 3000 ± 200 mg/L and 800 ± 100 mg/L) at θ_H anaerobic = 24 h on color and COD removal performance of the system were investigated. The results indicated that over 85% decolorization and about 90% COD removal efficiency can be obtained up to θ_H anaerobic = 48 h but higher retention times causes decreasing in decolorization efficiency. Operating the system with real wastewater without adding any nutrients at θ_H anaerobic = 24 h resulted in over 60% improvement in color removal in studied wastewater compared to existing treatment plant.