Bioremediation and biovalorisation of olive-mill wastes

Olive-mill wastes are produced by the industry of olive oil production, which is a very important economic activity, particularly for Spain, Italy and Greece, leading to a large environmental problem of current concern in the Mediterranean basin. There is as yet no accepted treatment method for all the wastes generated during olive oil production, mainly due to technical and economical limitations but also the scattered nature of olive mills across the Mediterranean basin. The production of virgin olive oil is expanding worldwide, which will lead to even larger amounts of olive-mill waste, unless new treatment and valorisation technologies are devised. These are encouraged by the trend of current environmental policies, which favour protocols that include valorisation of the waste. This makes biological treatments of particular interest. Thus, research into different biodegradation options for olive-mill wastes and the development of new bioremediation technologies and/or strategies, as well as the valorisation of microbial biotechnology, are all currently needed. This review, whilst presenting a general overview, focusses critically on the most significant recent advances in the various types of biological treatments, the bioremediation technology most commonly applied and the valorisation options, which together will form the pillar for future developments within this field.     

The Relationship Between the Metal Clusters and the Conformation of Molybdenum-Iron Protein from Azotobacter vinelandii

The ultraviolet CD spectrum of nitrogenase MoFe protein from Azotobacter vinelandii had a negative trough with double peaks at 208 nm and 222 nm, respectively, and the shape of the trough was similar to those of other proteins with a-helix structure. After treatment with o-phenanthroline under an aerobic or anaerobic condition, the height of the peak at 222 nm (h222 nm) decreased with the decrease of the C2H2-reduction activity, Fe content and CD spectra at both 450 nm and 660 nm, or at 450 nm of the treated proteins. However, after reconstituting with a reconstituent solution containing Na2MoO4, Na2S, dithiothreitol and either ferric homocitrate or ferric citrate, the h222 nm Of the reconstituted proteins could be restored as well as the activity, Fe content and CD spectra at both of 450 nm and 660 nm. The results show that there is a significant relationship between the metal clusters (FeMoco and P-cluster) and the conformation of MoFe protein.     

Effect of ammonium sulphate concentration and agitation speed on the kinetics of alginate production by Azotobacter vinelandii DSM576 in batch fermentation

Growth and alginate production by Azotobacter vinelandii DSM576 as a function of initial ammonium sulphate concentration (0.45–1.05 g l⁻¹) and agitation speed (300–700 rpm) were studied in batch fermentations at controlled pH. The time course of growth, alginate production and substrate consumption and the effect of nitrogen concentration and agitation speed on kinetic parameters and on maximum alginate molecular weight (MW) was modelled using empirical equations. The kinetics of growth, alginate production and polymerization were deeply affected by agitation speed and, to a lesser extent, by inorganic nitrogen concentration. Average and maximum specific growth rate and maximum alginate MW all increased with agitation speed, and were higher at intermediate ammonium sulphate concentration. Maximum alginate MW (>250,000) was obtained at high agitation speed (600–700 rpm) and alginate depolymerization was limited or did not occur at all when the agitation speed was higher than 500 rpm, while at 400 rpm depolymerization significantly reduced the alginate. However, alginate yield was negatively affected by increasing agitation speed. A good compromise between alginate yield (>2 g l⁻¹) and quality (MW>250,000) was obtained with agitation speed of 500–600 rpm and 0.75–0.90 g l⁻¹ of ammonium sulphate. Journal of Industrial Microbiology & Biotechnology (2000) 25, 242–248. Received 23 February 2000/ Accepted in revised form 04 August 2000     

Mutagenesis of a gene encoding a cytochrome o-like terminal oxidase of Azotobacter vinelandii: A cytochrome o mutant is aero-tolerant during nitrogen fixation

Abstract The amino acid sequence obtained by translating the nucleotide sequence of a 0.55 kb fragment, amplified from Azotobacter vinelandii chromosomal DNA by PCR, was 57% identical to part of the Escherichia coli cyoB gene, encoding subunit I of the cytochrome bo -type quinol oxidase. This fragment was mutated in vitro by insertion of a kanamycin-resistance cassette and introduced into the chromosome of A. vinelandii by homologous recombination. The mutant contained no spectrally detectable cytochrome o . However, in the stationary phase of growth, the level of the alternative oxidase (cytochrome bd ) was 11-fold higher than in the wild-type strain. Respiration of the mutant was insensitive to chlorpromazine, an inhibitor thought to act specifically on cytochrome o . Cytochrome o -deficient mutants fixed nitrogen in air, clearly distinguishing the role of this oxidase from that of cytochrome bd , which is required for respiratory protection of oxygen-labile nitrogenase.     

Cloning of the sth gene from Azotobacter vinelandii and construction of chimeric soluble pyridine nucleotide transhydrogenases

The gene encoding the soluble pyridine nucleotide transhydrogenase (STH) of Azotobacter vinelandii was cloned and sequenced. This is the third sth gene identified and further defines a new subfamily within the flavoprotein disulfide oxidoreductases. The three STHs identified all lack one of the redox active cysteines that are characteristic for this large family of enzymes, and instead they contain a conserved threonine residue at this position. The recombinant A. vinelandii enzyme was purified to homogeneity and shown to form filamentous structures different from those of Pseudomonas fluorescens and Escherichia coli STH. Chimeric STHs were constructed which showed that the C-terminal region is important for polymer formation. The A. vinelandii STH containing the C-terminal region of P. fluorescens or E. coli STH showed structures resembling those of the STH contributing the C-terminal portion of the protein.     

Relationship Between nifZ and the Synthesis of P cluster in Nitrogenase MoFe Protein of Azotobacter vinelandii

In comparison with OP MoFe protein from wild type strain Azotobacter vinelandii Lipmann, the C2H2-reduction activity and atom ratio of Fe to Mo of △nifZ MoFe protein from a nifZ deletion strain of A. vinelandii were remarkably decreased. FeMoco, which were extracted from these two proteins under the same condition, were almost similar to each other in activity and metal composition, and the circular dichroism (CD) spectra of these proteins were significantly different from each other. In the visible region except 540 750 nm, the △ε at 380 - 540 nm of △nifZ MoFe protein decreased and had a peculiar sharp negative peak around 430 nm; and in the ultraviolet region, the peaks at 208 nm and 222 nm were higher than those of OP MoFe protein. △nifZ MoFe protein could be crystallized in a suitable concentration of PEG 8000 and MgCl2, the size of crystals and amount of precipitation seemed to be related to the above-mentioned negative peaks. The results showed that △nifZ of Azotobacter vinelanclii might be related to the synthesis of P-cluster, rather than to that of FeMoco, which resulted in its conformation, stability and process of crystallization.     

Mutation of cytochrome bd quinol oxidase results in reduced stationary phase survival, iron deprivation, metal toxicity and oxidative stress in Azotobacter vinelandii

Azotobacter vinelandii cydAB mutants lacking cytochrome bd lost viability in stationary phase, irrespective of temperature, but microaerobiosis or iron addition to stationary phase cultures prevented viability loss. Growth on solid medium was inhibited by a diffusible factor from neighbouring cells, and by iron chelators, In(III) or Ga(III); microaerobic growth overcame inhibition by the extracellular factor. Siderophore production and total Fe(III)-chelating activity were not markedly affected in Cyd(-) mutants, and remained responsive to iron repression. Cyd(-) mutants were hypersensitive to Cu(II), Zn(II), and compounds exerting oxidative stress. Failure to synthesise haemoproteins does not explain the complex phenotype since mutants retained significant catalase activity. We hypothesise that Cyd(-) mutants are defective in maintaining the near-anoxic cytoplasm required for reductive iron metabolism and nitrogenase activity.     

Distributed state simulation of endogenous processes in biological wastewater treatment

Distributed state-type simulations (based on modeling of individual bacteria as they move through a reactor system) predicted a greater sensitivity of enhanced biological phosphorus removal (EBPR) performance to endogenous degradation than did conventional, "lumped"-type simulations (based on average biomass compositions). Recent research has indicated that the variable hydraulic residence times experienced by individual microbial storage product accumulating bacteria in systems with completely mixed reactors tend to produce populations with diverse microbial storage product contents (distributed states). Endogenous degradation in EBPR systems is of particular interest because the polyphosphate accumulating organisms (PAOs) responsible for EBPR rely on the accumulation of three different storage products that may be endogenously degraded. Simulations indicated that as endogenous degradation rates of microbial storage products were increased, EBPR performance decreased more rapidly according to the distributed approach than according to the lumped approach. State profile analysis demonstrated that as these rates increased, the population fraction with depleted storage products also increased, and this tended to increase the error in calculated biokinetic rates by the lumped approach. Simulations based on recently reported endogenous rate coefficients also suggested large differences between distributed and lumped predictions of EBPR performance. These results demonstrated that endogenous decay processes may play a more important role in EBPR than predicted by the lumped approach. This suggests a need for further research to determine endogenous process rates, and for incorporation of this information to distributed-type simulators, as this should lead to improved accuracy of EBPR simulations.     

Improvement strategy on enhanced biological phosphorus removal for municipal wastewater treatment plants: full-scale operating parameters, sludge activities, and microbial features

The poor quality of effluent discharged by municipal wastewater treatment plants (WWTPs) is threatening the safety of water ecology. This study, which integrated a field survey, batch tests, and microbial community identification, was designed to improve the effectiveness of the enhanced biological phosphorus removal (EBPR) process for WWTPs. Over two-thirds of the investigated WWTPs could not achieve total P in effluent lower than 0.5 mg/L, mainly due to the high ratio of chemical oxygen demand to P (28.6-196.2) in the influent. The rates of anaerobic P release and aerobic P uptake for the activated sludge varied from 0.22 to 7.9 mg/g VSS/h and 0.43 to 8.11 mg/g VSS/h, respectively. The fraction of Accumulibacter (PAOs: polyphosphate accumulating organisms) was 4.8 ± 2.0% of the total biomass, while Competibacter (GAOs: glycogen-accumulating organisms) accounted for 4.8 ± 6.4%. The anaerobic P-release rate was found to be an effective indicator of EBPR. Four classifications of the principal components were identified to improve the EBPR effluent quality and sludge activity.     

Effect of redox mediators on nitrogenase and hydrogenase activities in Azotobacter vinelandii

In bioelectrochemical studies, redox mediators such as methylene blue, natural red, and thionine are used to studying the redox characteristics of enzymes in the living cell. Here we show that nitrogenase activity in Azotobacter vinelandii is completely inhibited by oxidized methylene blue (MB_o) when the concentration of this mediator in the medium is increased up to 72 M. This activity in A. vinelandii is somewhat inhibited by a coenzyme, ascorbic acid (AA). However, the nitrogenase activity within the A. vinelandii cell is unchanged even for a high concentration of oxidized natural red (NR_o) alone. Interestingly, these mediators and AA do not have the capacity to inhibit the H₂ uptake activity of the hydrogenase in A. vinelandii. Average active rates of 66 nM H₂ evolved/mg cell protein/min from the nitrogenase and 160 nM H₂-uptake/mg cell protein/min from the hydrogenase in A. vinelandii are found in aid of the activities of the enzymes for H₂ evolution and for H₂ uptake are compared. The activities of both enzymes in A. vinelandii are strongly inhibited by thionine having high oxidative potential. Mechanisms of various mediators acting in vivo for both enzymes in A. vinelandii are discussed.     

Microbial population dynamics on seeds during drum and steeping priming

In the UK, seeds are primed commercially via drum or steeping priming processes to improve seed germination and seedling establishment but the microbial population dynamics occurring during these processes are unknown. Consequently, changes in culturable bacterial and fungal populations that occurred during laboratory and commercial scale drum priming of carrot, leek and parsnip seed and during laboratory scale and commercial scale steeping priming of carrot and sugar beet seed were investigated. Populations of bacteria and pseudomonads increased by 2 and 4 log₁₀ cfu g^–1 seed during priming to reach between 7 and 10 log₁₀ cfu g^–1 seed with less than 1 log₁₀ cfu g^–1 seed decrease after redrying, irrespective of seed type or priming process. Pseudomonads represented approximately 10% of the culturable bacterial population. Fungal populations also increased by between 1 and 3 log₁₀ cfu g^–1 seed during priming of carrot, leek and parsnip seed and were little affected by redrying. Addition of a tefluthrin coating (a standard commercial insecticide treatment) during the drying stage of commercial scale drum priming had little effect on microbial populations. During steeping priming, populations of bacteria and pseudomonads on sugar beet also increased by 2 and 4 log₁₀ cfu g^–1 seed with less than 1 log₁₀ cfu g^–1 seed decrease after redrying. However, fungi did not increase in the same manner on sugar beet, suggesting that the sugar beet seed itself may have an inhibitory effect on fungal growth. The presence of thiram in the steeping liquid (a standard commercial fungicide treatment) inhibited fungal proliferation on carrot and sugar beet seed. Spore forming bacteria generally occurred at low levels on all seeds (between 2 and 5 log₁₀ cfu g^–1seed) and did not exhibit any characteristic population changes during priming or redrying. Clearly some culturable populations of microorganisms can increase reproducibly during these priming processes and survive the redrying procedure, thereby demonstrating the potential for a novel method of introducing microbial inoculants onto seed.     

The macroecology of population dynamics: taxonomic and biogeographic patterns in population cycles

Regular cycles in population abundance are fascinating phenomena, but are they common in natural populations? How are they distributed among taxa? Are there differences between different regions of the world, or along latitudinal gradients? Using the new Global Population Dynamics Database we analysed nearly 700 long (25 + years) time series of animal field populations, looking for large-scale patterns in cycles. Nearly 30% of the time series were cyclic. Cycle incidence varied among taxonomic classes, being most common in mammal and fish populations, but only in fish did cycle incidence vary among orders. Cycles were equally common in European and North American populations, but were more common in Atlantic fish than Pacific fish. The incidence of cycles increased with latitude in mammals only. There was no latitudinal gradient in cycle period, but cycle amplitude declined with latitude in some groups of fish. Even after considering the biases in the data source and expected type I error, population cycles seem common enough to warrant ecological attention.     

Sexual reproduction and diversity: Connection between sexual selection and biological communities via population dynamics

Sexual reproduction is a mysterious phenomenon. Most animals and plants invest in sexual reproduction, even though it is more costly than asexual reproduction. Theoretical studies suggest that occasional or conditional use of sexual reproduction, involving facultative switching between sexual and asexual reproduction, is the optimal reproductive strategy. However, obligate sexual reproduction is common in nature. Recent studies suggest that the evolution of facultative sexual reproduction is prevented by males that coerce females into sexual fertilization; thus, sexual reproduction has the potential to enforce costs on a given species. Here, the effect of sex on biodiversity is explored by evaluating the reproductive costs arising from sex. Sex provides atypical selection pressure that favors traits that increase fertilization success, even at the expense of population growth rates, that is, sexual selection. The strength of sexual selection depends on the density of a given species. Sexual selection often causes strong negative effects on the population growth rates of species that occur at high density. Conversely, a species that reduces its density is released from this negative effect, and so increases its growth rate. Thus, this negative density-dependent effect on population growth that arises from sexual selection could be used to rescue endangered species from extinction, prevent the overgrowth of common species and promote the coexistence of competitive species. Recent publications on sexual reproduction provide several predictions related to the evolution of reproductive strategies, which is an important step toward integrating evolutionary dynamics, demographic dynamics and community dynamics.     

紫薇毡蚧种群生物学特性研究

该文报道紫薇毡蚧EriococcusLagerostroemiae Kuwana的种群生物学特性。该虫80年末传入贵州,在贵阳地区每年发生3-4代,世代重叠极重,越冬代是第3、4两代的混合群体,越冬虫态有卵、若虫和蛹。分别估测了卵、雄若虫、蛹、雌若虫和雌成虫的发育起点温度(T₀)和有效积温(K)。周年虫口变动有两个高峰：6月中旬至7月上旬和8月上旬至9月上旬。11月至翌年4月,因寄主休眠和低温导致虫口缓慢下降。雌成虫产卵量与寄主生长势好坏有关。控制紫薇毡蚧虫口最有效的天敌是红点唇瓢虫Chilocoruskuwanae Silverstri。药液涂干并辅之具内吸作用的杀虫药液灌根是目前较好的药剂防治方法。     

橄榄星室木虱的种群动态及药剂防治

橄榄星室木虱在福建省莆田地区 1年发生 8代 ,世代重叠 ,以成虫越冬 ,1年中有 6个发生高峰期 ,以橄榄秋梢期为全年雌成虫产卵、若虫发生最高峰 ,夏梢期是成虫羽化全年最高峰。田间药效试验结果表明 ,1 5种杀虫剂对橄榄星室木虱都有较好的防效 ,持效期可长达 7d以上 ,药后 7d的防效都在90 %以上 ,但速效性差异较大。其中 ,以 1 %威宝乳油 2 0 0 0倍、2 5 %功夫菊酯乳油 2 0 0 0倍、2 5 %功夫菊酯 +1 0 %吡虫啉 ( 1 2 5∶1 ) 2 0 0 0倍和 2 5 %功夫菊酯 +1 0 %吡虫啉 ( 1∶2 ) 2 0 0 0倍对橄榄星室木虱效果最好 ,速效性好 ,药后 1d的防效均达 91 5 8%以上。     

Microbial population dynamics and temperature changes during fermentation of kimjang kimchi

A distinct subset of lactic acid bacteria that are greatly influenced by temperature play an important role during kimchi fermentation. However, microbial population dynamics and temperature control during kimjang kimchi fermentation, which is traditionally fermented underground, are not known. Here we show that Lactobacillus sakei predominates in kimjang kimchi, perhaps due to suitable fermentation (5∼9°C) and storage (−2°C) temperatures. The temperature of this kimchi gradually decreased to 3.2°C during the first 20 days of fermentation (−0.3°C/day) and then was stably maintained around 1.6°C, indicating that this simple approach is very efficient both for fermentation and storage. These findings provide important information towards the development of temperature controlling systems for kimchi fermentation.