George A. Zavarzin

Alkaliphilic anaerobic community at pH 10

Relict or ancient microbial communities in extreme environment might be analogous to the centers of origin of bacterial diversity. A bacterial community of an alkaline lake was investigated, and the diversity of bacteria found there indicates that both conditions of autonomy and phylogenetic variety are fulfilled for anaerobic bacteria developing at pH 10±0.2. Major functional groups in the trophic network were present. Representatives of proteolytic, bacteriolytic, cellulolytic, saccharolytic, dissipotrophic, acetogenic, sulfate-reducing, methanogenic bacteria were isolated.

Emission of methane into the atmosphere from landfills in the former USSR

The annual production of solid domestic wastes by population of big cities of the USSR is about 37.5 million tons. The main method of disposal is burial in designated landfills. In the USSR large landfills occupy an area of more than 140,000 hectares. It has been calculated that the mass of the landfill deposits generating methane is today about 600 million tons. The studies carried out using geophysical, isotopic, and microbiological methods at different landfills of the Moscow region have shown that the emission of methane and other gases from the surface of landfills into the atmosphere is extremely irregular and considerably less than their generation in the anaerobic zone. The most important factors determining methane emission are the thickness of the layer of buried refuse, the heterogeneity of the deposit body, and the microbiological oxidation of gases in the upper aerated ground layer. It has been shown that the temperature in the anaerobic zone of big landfills is relatively constant and in most cases is 25–35°C. Methanogenesis often is most intensive in the upper part of anaerobic zone where the content of organic matter is rather high. The stable carbon isotope composition of the biogas generated from landfills is characteristic of the methanogenesis from organic wastes and depends on the concentration of organic matter and the age of the landfill. At first a lighter gas is generated and then a heavier one as the substrate is depleted. In the upper aerated ground layer of the landfill, about 1 meter in depth, methane, hydrogen, and carbon monoxide (CH4, H2, CO) are oxidized intensively. The number of bacteria oxidizing these gases reaches 1011 cells per gram of wet refuse. In this case the stable isotope composition of methane becomes heavier and, of carbon dioxide, lighter. It has been shown that at small landfills methane can be oxidized completely in aerobic zones. The gas-oxidizing ability of the microflora of the aerated ground layer of a landfill decrease considerably in the cold season of the year. The methane emission from landfills located in the USSR is estimated at 1.2 – 2.4 billion cubic meters per year. About two-thirds comes from the European part of the country. Effective methods of decreasing methane emission into the atmosphere are the extraction of biogas from big landfills and the maintenance of good aeration of the upper ground layer at the small ones.

Halocella cellulolytica gen. nov., sp. nov., a New Obligately Anaerobic, Halophilic, Cellulolytic bacterium

Summary A new halophilic cellulolytic bacterium isolated from hypersaline lagoons of Lake Sivash (Crimea) is described. The organism is an obligate anaerobe. Concentrations of between 5% and 20% NaCl are required for growth; optimum growth occurs at a concentration of 15% NaCl. The cells are Gram-negative, motile rods, 0.4–0.6 × 3.8–12.0 μm in size. Cellulose, starch and a wide variety of carbohydrates are utilized. The products of cellulose fermentation are acetate, ethanol, lactate, hydrogen and carbon dioxide. The temperature range for growth is 20–50°C (opt. 39°C). The pH range is 5.5–8.5 (opt. 7.0). The G+C content of the DNA is 29 mol.%. The bacterium differs from other anaerobes and halophiles described in the literature. The name Halocella cellulolytica gen. nov., sp. nov. is proposed. The type strain is z-10151.

Tindallia magadii gen. nov., sp. nov.: An Alkaliphilic Anaerobic Ammonifier from Soda Lake Deposits

Abstract. Strain Z-7934, an alkaliphilic, obligately anaerobic, fermentative, asporogenous bacterium with Gram-positive cell wall structure, was isolated from soda deposits in Lake Magadi, Kenya. The organism ferments only a few amino acids, preferentially arginine and ornithine, with production of acetate, propionate, and ammonia. It is a true alkaliphile, with pH range for growth ranging from 7.5 to 10.5 (optimum pH 8.5), and growth is dependent on the presence of sodium ions. The G+C content of the genomic DNA is 37.6 mol%. 16S rDNA sequence analysis of strain Z-7934 shows that it belongs phylogenetically to cluster XI of the low G+C Gram-positive bacteria. On the basis of its distinct phylogenetic position and unique physiological properties, we propose a new genus and new species, Tindallia magadii, for this strain. The type strain is Z-7934T (=DSM 10318).

Spirochaeta thermophila sp. nov., an Obligately Anaerobic, Polysaccharolytic, Extremely Thermophilic Bacterium

Growth at temperatures of >60°C and utilization of polysaccharides have not been reported previously in members of the genus Spirochaeta. Two obligately anaerobic, extremely thermophilic (optimum temperature, 65°C) spirochetes were isolated from geographically distant thermal sites. These two isolates have chemoorganotrophic fermentative metabolism and grow on a variety of mono-, di-, and polysaccharides, including cellulose. The differences in pH and NaCl concentration optima between these organisms reflect the prevailing conditions at the sites from which they were isolated. DNA-DNA hybridization showed that the two strains exhibit a level of homology of 87%. On the basis of their morphological characteristics, their high level of homology with each other, and their extremely thermophilic and polysaccharolytic nature, we propose that these organisms should be included in the genus Spirochaeta as a new species, Spirochaeta thermophila; the type strain of this species is strain Z-1203 (= DSM 6578).

Ecology, Physiology and Taxonomy Studies on a New Taxon of Haloanaerobiaceae, Haloincola saccharolytica gen. nov., sp. nov.

Summary Four strains of new moderately halophilic, Gram-negative anaerobic saccharolytic bacteria were isolated from Lake Sivash and its lagoons on Arabat Strait (eastern Crimea, USSR). The cells are non-sporeforming, motile rods 0.5-0.7 × 1-1.5 μm, that occur single or in pairs. Carbohydrates (preferably disaccharides, such as sucrose and trehalose) but not amino acids serve as energy sources. The end products of glucose fermentation are acetate, H 2 and CO 2 . The biomass yield is 3.8 g of protein per mol of glucose consumed. Elemental sulphur when present is reduced to H 2 S. All isolates are mesophiles and grow at salt concentrations from 3 to 30% NaCl (w/v) and at a pH range from 6.0 to 8.0. Doubling time of strain Z-7787 at optimal conditions (37°C., 10% NaCl, pH 7.5) is 3.9 h. The G+C content of this isolate is 31.3 mol%; that of the other strains varies from 30 to 33 mol%. All the isolates appear to be members of the same taxon but according to 5S rRNA sequencing do not belong to the genus Halobacteroides . They are considered to represent a new genus for which the name Haloincola is proposed with Haloincola saccharolytica as the type species. The type strain is Z-7787, DSM 6643. The taxonomic relationships of H. saccharolytica to other haloanaerobes of the family Haloanaerobiaceae is discussed.

Halophilic Acetogenic Bacteria

As outlined in preceding chapters of this book, acetogenic bacteria have a specialized physiological potential for the conservation of energy via the reduction of CO2 to acetate. They also harbor diverse catabolic processes and are found in unusual habitats. Although their role in nature was initially viewed somewhat restrictively, it is now evident that they might have a large impact on carbon and energy flow in some environments, in particular that of certain gastrointestinal ecosystems (Breznak et al., 1988; Kane and Breznak, 1991a, 1991b). However, specialized (or extreme) terrestrial environments are largely unexplored relative to the involvement of acetogenesis and associated organisms.

Phylogenetic Study of Haloanaerobic Bacteria by 16S Ribosomal RNA Sequences Analysis

Summary The nucleotide sequences of 16S rRNA of four strains of the haloanaerobic bacteria Haloincola saccharolytica Acetohalobium arabaticum, Halobacteroides lacunaris, and Halobacteroides sp. have been determined. The phylogenetic analysis employing the MTP method placed the group of the haloanaerobes in a single cluster at the base of a branch leading to the Gram-positive organisms on the constructed unrooted tree. However, an additional analysis demonstrated that the cluster of the haloanaerobes did not belong to any subdivision of Gram-positive bacteria, including the representatives of the Gram-negative branches — Heliobacterium and Sporomusa. The signature analysis and the secondary structure analysis of the 16S rRNA of the haloanaerobes revealed some peculiarities that distinguish them from the other bacterial phyla. All these data along with the unique phenotype of these microorganisms demonstrate that the haloanaerobes represent a separate line of descent within the Bacteria.

Methanethiol utilization and sulfur reduction by anaerobic halophilic saccharolytic bacteria

A number of sulfur compounds were tested as sulfur sources for the growth of three strains of anaerobic halophilic saccharolytic bacteria isolated from hypersaline water bodies of the eastern Crimea (USSR). Dithionite and sulfite at 1 mM concentration completely inhibited the growth of all strains. Methanethiol turned out to be the sole sulfur source for growth ofHalobacteroides strains in the defined medium with glucose and leucine. Methanethiol also stimulated growth of cultures in the complex medium with yeast extract. TheHaloincola saccharolytica Z-7787 appeared to be capable of methanethiol formation from methionine. All organisms studied were capable of heterotrophic sulfur reduction, producing up to 13 mM H2S, but no evidence that they gain energy from the process has been obtained. The extremely halophilicHalobacteroides lacumaris may participate in sulfidogenesis at the high salinity (20–30% NaCl). The ecological position of haloanaerobes in halophilic community is discussed.

Landfills and waste deposits producing greenhouse gases

Abstract Man-made landscapes may be integrated into the natural environment by new combinations of conventional microbial processes. Landfills and waste deposits are extreme examples of artificial landscapes, but their influence on the biosphere is only minimal because they are analogous to geological sediments and they harbour a microbial community that catalyzes closed cycles. Greenhouse gas production from landfills is of global significance while contamination of ground water is of greater importance on a local scale.