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.     

Methanocalculus natronophilus sp. nov., a new alkaliphilic hydrogenotrophic methanogenic archaeon from a soda lake,and proposal of the new family Methanocalculaceae

A mesophilic hydrogenotrophic methanogenic archaeon, strain Z-7105^T, was isolated from the bottom sediments of a collector in the vicinity of a soda lake Tanatar II (Altai, Russia). The cells were motile, irregular cocci 0.2–1.2 μm in diameter. The organism was an obligate alkaliphile, growing within a pH range from 8.0 to 10.2, with the optimum at pH 9.0–9.5. It was obligately dependent on carbonates, growing at 0.5 to 1.6 M total carbonates with the optimum at 0.7–0.9 M. Sodium ions were also obligately required at concentrations from 0.9 to 3.3 M Na⁺ (optimum at 1.4–1.9 M). The organism was halotolerant, but Clions were not required. Hydrogen and formate were used as electron donors. Acetate was required for anabolism. The DNA G+C content was 50.2 mol %. According to the results of its 16S rRNA gene sequence analysis, the isolate belonged to the genus Methanocalculus, being the first known alkaliphilic member of this genus. Its similarity to the neutrophilic and halotolerant Methanocalculus species (M. halotolerans, M. taiwanensis, M. pumilus, and M. chunghsingensis) was 98.2–97.1%, which is within the interspecific range for this genus. The level of DNA-DNA hybridization between strain Z-7105^T and the Methanocalculus type species M. halotolerans DSM 14092^T was 32%. The genus Methanocalculus, including the new isolate and the previously described species, is distant from other genera of methanogens (<90% 16S rRNA gene similarity). Based on significant phenotypic differences and the results of phylogenetic analysis, including DNA-DNA hybridization, it is proposed to assign strain Z-7105^T (=DSM 25006^T, =VKM B-2765^T) to the new species Methanocalculus natronophilus sp. nov., and to incorporate the genus into the new family Methanocalculaceae fam. nov.     

Reduction of synthetic ferrihydrite by a binary anaerobic culture of <Emphasis Type="Italic">Anaerobacillus alkalilacustris</Emphasis> and <Emphasis Type="Italic">Geoalkalibacter ferrihydriticus</Emphasis> grown on mannitol at pH 9.5

In the course of an investigation of alkaliphilic iron reduction, metabiotic interactions in a binary culture reducing synthetic ferrihydrite (SF) have been studied. The binary culture contained two anaerobic bacteria: the alkaliphilic organotrophic bacillus Anaerobacillus alkalilacustris, which ferments sugars and sugar alcohols and is incapable of iron reduction, and the dissimilatory iron-reducing bacterium Geoalkalibacter ferrihydriticus, which is able to grow on acetate at the expense of anaerobic respiration. The experiments were performed under conditions of SF excess and deficiency. It was expected that G. ferrihydriticus would oxidize the acetate formed in the course of mannitol fermentation by A. alkalilacustris. The results were different from the expected ones: in the binary culture, fermentation products other than acetate were used for iron reduction; these were primarily formate and ethanol, which led to acetate accumulation rather than consumption. The reduction of SF to magnetite and/or siderite followed the earlier established regularities. The preferential order of donor utilization by G. ferrihydriticus did not conform to the energy yields of the corresponding reactions. Thus, it has been shown that there may be interactions in microbial communities that cannot be predicted from the characteristics of pure cultures. The degradation pathways of organic matter in communities may differ considerably from those observed in pure cultures, even in pure cultures of highly specialized organisms.     

Cellulase activity of a haloalkaliphilic anaerobic bacterium, strain Z-7026

Summary The cellulolytic activity of an alkaliphilic obligate anaerobic bacterium, Z-7026, which was isolated from the microbial community of soda-lake sediments and belongs to the cluster III of Clostridia with low G+C content, was studied. The bacterium was capable of growing in media with cellulose or cellobiose as the sole energy sources. Its maximal growth rate on cellobiose (0.042–0.046 h^–1) was observed at an initial pH value of 8.5–9.0, whereas the maximal rate of cellulase synthesis, assayed by using a novel fluorimetric approach, was found to be 0.1 h^–1 at pH 8–8.5. Secreted proteins revealed high affinity for cellulose and were represented by two major forms of molecular masses of 75 and 84 kDa, whereas the general protein composition of the precipitated and cellulose-bound preparations was similar to cellulosome subunits of Clostridium thermocellum. The optimum pH of the partially purified enzyme preparation towards both amorphous and crystalline cellulose was in the range 6–9, with more than 70% and less than 50% of maximal activity being retained at pH 9.2 and 5.0, respectively.     

DNA relatedness among some thermophilic members of the genus Methanobacterium: emendation of the species Methanobacterium thermoautotrophicum and rejection of Methanobacterium thermoformicicum as a synonym of Methanobacterium thermoautotrophicum.

DNA reassociation was used to determine levels of relatedness among four thermophilic Methanobacterium strains that are able to use formate and between these organisms and two representative strains of Methanobacterium thermoautotrophicum, strain delta HT (= DSM 1053T = ATCC 29096T) (T = type strain) and strain Marburg (= DSM 2133). Three homology groups were delineated, and these groups coincided with the clusters identified by antigenic fingerprinting. The first group, which had levels of cross hybridization that ranged from 73 to 99%, included M. thermoautotrophicum delta HT, Methanobacterium thermoformicicum Z-245, Methanobacterium sp. strain THF, and Methanobacterium sp. strain FTF. The second and third groups were each represented by only one strain, Methanobacterium sp. strain CB-12 and M. thermoautotrophicum Marburg, respectively (cross-hybridization levels, 13 to 30 and 29 to 33%, respectively). Our results indicate that the name M. thermoformicicum should be rejected as it is a synonym of M. thermoautotrophicum. The taxonomic positions of strains Marburg and CB-12 need further investigation.     

Proteinivorax tanatarense gen. nov., sp. nov., an anaerobic, haloalkaliphilic, proteolytic bacterium isolated from a decaying algal bloom, and proposal of Proteinivoraceae fam. nov.

Two strains of a novel anaerobic, protein- and nucleoside-utilizing bacterium, Z-910^T and Z-810, were isolated. The strains were spore-forming, mainly nonmotile rods, exhibiting positive Gram reaction with Gram-positive cell wall structure. The strains were mesophilic and haloalkaliphilic. Cultures used proteins and proteinaceous substrates as carbon, nitrogen, and energy sources. Both strains used also ribonucleosides, cellobiose, pyruvate, and glycerol. Ribose and nucleobases did not support growth. The fermentation products from all utilized substrates were identical but varied in content and included straight and branched acids, as well as hydrogen and ammonia. When grown on tryptone, strain Z-910^T was able to reduce fumarate, dimethyl sulfoxide, thiosulfate, and elemental sulfur. Neither nitrate nor sulfate was reduced. The DNA G + C content of strain Z-910^T was 32.2 mol%. Phylogenetic analysis based on the 16S rRNA gene sequence similarity revealed that strains Z-910^T and Z-810 represented a new branch within the order Clostridiales, with 90.2 % similarity to the nearest genus with a validly published name Anaerobranca gottschalkii DSM 13577^T. According to their physiological, chemotaxonomic, and phylogenetic properties, strains Z-910^T and Z-810 represented a new genus and novel species, for which the name Proteinivorax tanatarense gen. nov., sp. nov. was proposed. Phylogenetic analysis showed that the genera Proteinivorax gen. nov. and Anaerobranca formed a separate cluster within the order Clostridiales. The family Proteinivoraceae fam. nov. comprising the genera Proteinivorax gen. nov. and Anaerobranca was therefore proposed within the order Clostridiales of the phylum Firmicutes with Proteinivorax as a type genus of the new family.