Trello Beffa

Taxonomic and Metabolic Microbial Diversity During Composting

A great variety and high numbers of aerobic thermophilic heterotrophic and/or autotrophic bacteria growing at temperatures between 60–80°C have been isolated from thermogenic (temperature 60–80°C) composts in several composting facilities in Switzerland. They include strains related to the genus Thermus (T. thermophilus. T. aquaticlls. and several other new strains). Bacillus schlegelii, Hydrogenohacter spp., and of course heterotrophic sporeforming Bacilli. This contrasts with the generally held belief that thermogenic composts (> 60°C) support only a very low diversity of heterotrophic thermophiles. This biodiversity suggests efficient decomposition of organic matter at temperatures above 60°C, and a good thermo-hygienization.

Obligately and facultatively autotrophic, sulfur- and hydrogen-oxidizing thermophilic bacteria isolated from hot composts

Abstract A variety of autotrophic, sulfur- and hydrogen-oxidizing thermophilic bacteria were isolated from thermogenic composts at temperatures of 60–80° C. All were penicillin G sensitive, which proves that they belong to the Bacteria domain. The obligately autotrophic, non-spore-forming strains were gram-negative rods growing at 60–80°C, with an optimum at 70–75°C, but only under microaerophilic conditions (5 kPa oxygen). These strains had similar DNA G+C content (34.7–37.6 mol%) and showed a high DNA:DNA homology (70–87%) with Hydrogenobacter strains isolated from geothermal areas. The facultatively autotrophic strains isolated from hot composts were gram-variable rods that formed spherical and terminal endospores, except for one strain. The strains grew at 55–75° C, with an optimum at 65–70° C. These bacteria were able to grow heterotrophically, or autotrophically with hydrogen; however, they oxidized thiosulfate under mixotrophic growth conditions (e.g. pyruvate or hydrogen plus thiosulfate). These strains had similar DNA G+C content (60–64 mol%) to and high DNA:DNA homology (> 75%) with the reference strain of Bacillus schlegelii. This is the first report of thermogenic composts as habitats of thermophilic sulfur- and hydrogen-oxidizing bacteria, which to date have been known only from geothermal manifestations. This contrasts with the generally held belief that thermogenic composts at temperatures above 60° C support only a very low diversity of obligatory heterotrophic thermophiles related to Bacillus stearothermophilus.

Aspergillus fumigatus in windrow composting : effect of turning frequency

The influence of the windrow turning frequency on the pro liferation and dispersion of Aspergillus fumigatus (AF) at a classic, open-air windrow composting site was investigated. Independent of the turning frequency, the intially high con centrations of AF (>106 cfu/gDW ) were strongly reduced already after 2 weeks of composting in the center of all exper imental windrows, as well as at the surface of those being turned frequently (daily). Surface AF counts, however, remained elevated (>10 4 cfu/gDW ) for the less frequently (weekly to monthly) turned windrows. Also, in this case, con centrations of AF dispersed in the air were higher. Tempera ture, the main factor for eliminating AF, rose faster, more homogeneously, and reached higher values in the more fre quently (daily and/or weekly) turned composts than in those turned less frequently. Significant differences were seen between daily and weekly turnings of material with high a C/N ratio (40: 1), but not for the more nutrient rich composts (C/N 30:1), where temperatures were generally higher.

Linear growth and poly(β-hydroxybutyrate) synthesis in response to pulse-wise addition of the growth-limiting substrate to steady-state heterotrophic continuous cultures of Aquaspirillum autotrophicum

Heterotrophic pyruvate-limited steady-state continuous cultures of the bacterium Aquaspirillum autotrophicum were perturbed with a pulse injection of a small volume of concentrated pyruvate solution. These cultures exhibited an instantaneous change in the growth dynamics, turning from steady state to apparently linear growth. These transient growth-responses had no lag phase and were clearly distinct from unlimited exponential growth according to the initial rates of increase of biomass and substrate disappearance kinetics. A linear accumulation with time of poly(beta-hydroxybutyrate) was observed within the cells. Slopes of these linear responses were negatively correlated with the dilution rate. Physiological bases of linear growth are discussed in the light of the models of H. E. Kubitschek. Poly(beta-hydroxybutyrate) synthesis in the absence of exogenous limitation may serve to protect the cells against a transient metabolic overflow.

Growth and respiratory oxidation of reduced sulfur compounds by intact cells ofThiobacillus novellus (type strain) grown on thiosulfate

Thiobacillus novellus (type strain) was grown chemolithoautrophically on thiosulfate in batch cultures under microaerophilic conditions. Under these conditions,T. novellus grew exponentially (μ=0.05–0.06 h−1). The respiratory oxidation rates of tetrathionate, thiosulfate, elemental sulfur (So), and sulfite were measured respirometrically with an oxygen electrode, with exponentially growing cells. Cells growing on thiosulfate as the unique energy source retain thiosulfate-oxidizing activity, So-oxidizing activity (SOA), and very high sulfite-oxidizing activity, but lack respiratory tetrathionate-oxidizing activity. HQNO (50 μm), an inhibitor of the quinone-cytochrome b region, strongly inhibited the SOA (70%), moderately the sulfite-oxidizing activity (45%), and poorly the thiosulfate-oxidizing activity (15%), 1mm KCN totally inhibited (>89%) all respiratory activities. This study confirms that inThiobacillus novellus, as well as in otherThiobacilli, SOA is present in cells grown with thiosulfate as sole electron donor. SOA appears not to be an oxygenase; it is linked to the respiratory chain, and the electrons are probably released in the quinone-cytochrome b region.

Respiratory oxidation of reduced sulfur compounds by intact cells of Thiobacillus tepidarius (type strain)

Thiobacillus tepidarius (type strain) was grown in microaerophilic conditions, on tetrathionate, thiosulfate or crystalline So. The rates of tetrathionate, thiosulfate, elemental sulfur (So) and sulfite oxidation of the different cultures were measured respirometrically, using exponentially growing cells, with an oxygen electrode. Cells growing on the three different sulfur compounds retain thiosulfate-, tetrathionate, and So-oxidizing activities (SOA), but lack respiratory sulfite-oxidizing activity. The SOA for all the cultures was almost totally inhibited by 50 μM myxothiazol, an inhibitor of the quinone-cytochrome b region, and by 10 μM of the uncoupler carbonyl cyanide m-chlorophenylhydrazone (CCCP). Tetrathionate- and thiosulfate-oxidizing activities were moderately and weakly inhibited by 50 μM totally inhibited (>95%) all respiratory activities. This study suggests that electrons released by So oxidation enter the respiratory chain in the quinone-cytochrome b region.

Elemental sulfur production during mixotrophic growth on hydrogen and thiosulfate of thermophilic hydrogen-oxidizing bacteria

In this study we measured the exogenous production and the intracellular content of elemental sulfur (S0) in the thermophilic sulfur-oxidizing bacteriaHydrogenobacter spp. andBacillus schlegelii during mixolithoautotrophic growth on hydrogen and thiosulfate. Under these conditions, all strains produced and released white-yellow hydrophilic S0 particles into the growth medium. Hydrophilic S0 was separated from cells by a differential low-speed centrifugation procedure. The S0 pellets were dried, and the S0 was purified by column chromatography and by thin-layer chromatography (TLC). The S0 TLC-band could be stained with triphenyltetrazolium chloride and piperidine procedure. Determination of intracellular S0 content was performed from fresh cells absolutely free of exogenous S0 particles. quantitation of S0 was performed by high-performance liquid chromatography, colorimetric thiocyanate procedure, and by UV-spectra analyses. All the strains studied, in particularB. schlegelii strains, released significant quantities of S0 into the growth media. In contrast, the intracellular S0 content was very low. Significant rhodanese activity in the presence of thiosulfate was measured with toluenepermeabilized cells and cell-free extracts ofB. schlegelii (type strain) andHydrogenobacter spp. (strain T3).

Composting of Organic Garden and Kitchen Waste in Open-air Windrows: Influence of Turning Frequency on the Development of Aspergillus Fumigatus

Composting at industrial scale (installations treating. 100 tons per year) is increasingly used in Switzerland for the stabilization of organic waste and the recycling of humigenic materials. In 1992, 320.000 tons of garden and kitchen waste were treated in 150 composting sites, 80% thereof treating more than 1.000 t/a (1). Even if mature composts are generally of satisfactory quality, the process itself could often be considerably improved. Our research team is currently involved in a threeyear study that aims at optimizing the thermogenic phase of composting processes, in order to ensure a good thermo-hygienization (elimination of allergenic and pathogenic micro-organisms, e.g. the thermotolerant mold Aspergillus fumigatus) and to improve the number and diversity of the thermophilic micro-organisms. This will lead to a rapid degradation of the organic material. By this, phytotoxicity hazards can also be avoided.

Endogenous elemental sulfur production froml-cysteine in dormant α-spores ofPhomopsis viticola

The enzymatic production of sulfur froml-cysteine was studied in young dormant α-spores ofPhomopsis viticola. Cysteine aminotransferase (CAT) and mercaptopyruvate sulfurtransferase (MST) activities could be responsible for the production of endogenous elemental sulfur (S0) in α-spores.l-Cysteine was first deaminated, with production of β-mercaptopyruvate, by the CAT. The β-mercaptopyruvate produced is successively desulfurated by the MST with production of sulfur and pyruvate. Deaminase activity was recovered principally in the cytoplasmic fraction, whereas desulfurase activity was recovered mainly in the mitochondrial fraction.l-Cysteine and S0 sharply affected the respiratory activity, the ATP content, and suppressed germination of α-spores. In contrast, reduced glutathione did not affect these metabolic parameters. Production of S0 by enzymatic degradation ofl-cysteine could be responsible for the inhibitory action of this amino acid. We suggest that CAT and MST, by their capacity to produce sulfur or S0, plays a key role in regulation of morphogenetic processes ofPhomopsis viticola.

Biodiversity of Thermophilic Bacteria Isolated from Hot Compost piles

Composting is a self-heating, aerobic solid phase biodegradative process of organic waste materials, making possible its return to the environment as soil fertilizer and conditioner (Finstein & Morris, 1975; de Bertoldi & Zucconi, 1987; Finstein, 1992). Temperature increase involves a rapid transition from a mesophilic to a thermophilic microflora (65–75 °C) within a few hours, providing the pile is regularly aerated or frequently turned.