The peptidase activity of the alkaliphilic, aerobic, proteolytic bacteria Alkalicaulis satelles G-192T and Aliidiomarina sp. P-156 isolated from the system of hypersaline, alkaline Tanatar lakes (Altai Territory) was studied. Strains G-192 and P-156 710 were shown to hydrolyze para-nitroanilide substrates and exhibit the highest activity hydrolyzing of the aminopeptidase LpNa substrate. Analysis of partially purified peptidase preparations showed that the enzymes were most active and stablest in an alkaline pH range of 8.4-11. The peptidases of strains G-192 and P-156 were highly stable in NaCl up to 220 and 70 g/L respectively. The results of inhibitor analysis and the substrate specificity of the studied extracellular enzymes indicated their classification as metallopeptidases of the aminopeptidase type. The studied peptidases showed significant resistance to the surfactants Triton X-100 and SDS and the oxidizing agent H2O2. The isolated bacteria that produce peptidases can be used as a source of proteolytic enzymes in the development of new detergents.

A prosthecate bacterial strain, designated G-192T, was isolated from decaying biomass of a haloalkaliphilic cyanobacterium Geitlerinema sp. Z- T0701. The cells were aerobic, Gram- negative, non- endospore- forming and dimorphic, occurring either as sessile bacteria with a characteristic stalk or as motile flagellated cells. The strain utilized a limited range of substrates, mostly peptonaceous, but was able to degrade whole proteins. Growth occurred at 5?46 ?C (optimum, 35?40 ?C), pH 7.3?10.3 (optimum, pH 8.0?9.0), 0?14 % NaCl (v/w; optimum, 2.0?6.0 %, v/w). The G+C content of the genomic DNA of strain G-192T was 66.8%. Phylogenetic analysis of the 16S rRNA gene sequence revealed that strain G-192T formed a distinct evolutionary lineage within the family Hyphomonadaceae. Strain G-192T showed the highest 16S rRNA sequence similarity to Glycocaulis profundi ZYF765T (95.2%), Oceanicaulis stylophorae GISW-4T (94.2%) and Marinicauda salina WD6-1T (95.5%). The major cellular fatty acids (>5% of the total) were C18:1?9c, C18:0 and 11-methyl-C18:1 ?7c. The major polar lipids were glycolipids and phospholipids. The only respiratory quinone was ubiquinone-10 (Q-10). Based on polyphasic results including phylogenomic data, the novel strain could be distinguished from other genera, which suggests that strain G-192T represents a novel species of a new genus, for which the name Alkalicaulis satelles gen. nov., sp. nov. is proposed. The type strain is G-192T (=VKM B-3306T=KCTC 72746T). The strain is the first representative of the stalked bacteria associated with a haloalkaliphilic cyanobacterium. Based on phylogenomic indices and phenotypic data, it is proposed to evolve two novel families Maricaulaceae fam. nov. and Robiginitomaculaceae fam. nov. out of the current family Hyphomonadaceae. In addition, it is proposed to place the first two families in the novel order Maricaulales ord. nov. and novel order Hyphomonadales ord. nov. is proposed to accommodate the family Hyphomonadaceae.

A novel haloalkaliphilic bacterium, designated G-116(T), was isolated from the decaying biomass of a laboratory culture of cyanobacterium Geitlerinema species. Cells of strain G-116(T) were Gram-stain-negative, motile spirilla. Strain G-116(T) showed high halotolerance to 20 % (w/v) NaCl (optimum growth at 3.5-6.0 %, w/v) and obligately alkaliphilic growth within the pH range 7.3-10.4 (optimum growth at pH 8.7-8.9). The major fatty acids identified were C-16:0, summed feature 8 (C-18:1 omega 7c/C-18:1 omega 6c), summed feature 3 (C-16:1 omega 7c/C-16:1 omega 6c) and C-19:0 cyclo omega 8c. The polar lipids consisted of phosphatidylethanolamine, phosphatidylglycerol, diphosphatidylglycerol, an unidentified phospholipid, three unidentified amino lipids and five unidentified lipids. The predominant respiratory quinone was ubiquinone-8 (Q-8). The G+C content of the genomic DNA was 60.4 mol%. Phylogenetic analysis based on 16S rRNA gene sequences revealed that the closest genus with a validly published name is a monotypic Salinispirillum and strain G-116(T) clustered with Salinispirillum marinum GCWy1(T) with a 16S rRNA gene sequence similarity of 94.3 %. Based on the data obtained from phenotypic and chemotaxonomic studies and the phylogenetic analysis, the isolate is proposed to be a representative of a novel genus and a novel species, Natronospirillum operosum gen. nov., sp. nov. Together with S. marinum they form a separate clade, for which a novel family, Natronospirillaceae fam. nov., is proposed. In addition, Saccharospirillaceae fam. nov. and Gynuellaceae fam. nov. are proposed to encompass the genera Saccharospirillum and Reinekea, and the genus Gynuella, respectively. All three novel families are within the order Oceanospirillales of the class Gammaproteobacteria. The type strain of the type species, Natronospirillum operosum gen. nov., sp. nov. is G-116(T) (=VKM B-3134(T)=KCTC 62956(T)).

Alkaliphilic microorganisms are ubiquitous and inhabit various econiches on Earth. Alkaline environments suitable for alkaliphilic microbial communities may be created by certain geological processes or human activities. Moreover, a significant contribution to the emergence of alkaline conditions may be due to the activity of neutralophilic microorganisms through certain reactions, which explains the widespread distribution of alkaliphiles. Alkaliphilic microorganisms are part of extremophiles and become interesting and useful in environmental and industrial microbiology. With increasing knowledge of alkaliphiles, we greatly increase their biotechnological and industrial application potential. New microorganisms from natural habitats serve as a practically endless source of new enzymes. This chapter summarizes the scattered data on alkaliphiles isolated from habitats other than soda lakes. The conditions for occurrence of alkaline habitats are also considered. Moreover, the chapter reviews some important features on preparation of media for the isolation and cultivation of alkaliphiles. The chapter also includes relevant information on chromatographic analysis of alkaliphiles culture.

Peptidase activity of alkaliphilic anaerobic proteolytic Proteinivorax bacteria, P. tanatarense Z-910(T) and P. hydrogeniformans Z-710(T), isolated from the Tanatar system of hypersaline alkaline lakes (Altai krai, Russia) was studied. Strains Z-910 and Z-710 were shown to hydrolyze para-nitroanilide substrates, exhibiting the highest activity hydrolyzing GlpAALpNA and LpNa. The peptidases were most active under alkaline conditions at pH 8-10 and were stable at temperatures of up to 50 degrees C. The results on inhibitor analysis and substrate specificity of the studied extracellular enzymes suggested their classification as serine peptidases of the subtilisin-like type (strain Z-910) and metallopeptidases of the aminopeptidase type (strain Z-710).

The work continues the series of long-term studies of the microbial diversity of soda lakes and concerns the problem of anaerobic decomposition of proteins. The physiology of new proteolytic bacteria has been investigated. The lytic effect of the alkaliphilic anaerobe Proteinivorax tanatarense on other members of community, primarily cyanobacteria, has been demonstrated. P. tanatarense proved to be a saprotrophic satellite regulating the number of cyanobacteria under night conditions. The directed effects of P. tanatarense on phototrophic and chemotrophic Gram-negative microorganisms were demonstrated as well as the absence of its effect on any Gram-positive microorganisms. Such specialization of proteolytic agent could be implemented in cyanobacterial communities of the past.