Polyphase Taxonomy of Antarctic Bacteria

Abstract

The phylogenetic structure of bacteria is not always consistent with the traditional classification scheme based on the phenotypic properties of bacteria. That is one of the problems of modern bacterial taxonomy. In addition, traditional methods to identify bacteria using phenotypic properties have a number of disadvantages. In recent decades, significant progress was achieved in the investigation of microbial world using molecular methods for fast identification. The aim of the study was to clarify the species status of four strains of bacteria isolated from black lichens of the cliffs of Galindez Island in the Antarctic on the basis of phenotypic and genetic analysis. Methods. Morphological and cultural properties of bacteria were studied according to generally accepted microbiological methods. Physiological and biochemical ones were investigated using test systems API Coryne and API 20E (bioMerieux SA, France), according to the manufacturer’s instructions. Phylogenetic analysis was performed based on the nucleotide sequences of the 16S rRNA gene. To identify closely related species, a comparative analysis of the nucleotide sequences of 16S rRNA genes was performed using the BLAST software package. The phylogenetic position was determined by constructing trees (dendrograms) to show the position of the studied strains among closely related and typical species (programs ClustalX 2.1, Mega 6.06). The tree was constructed applying ClustalX 2.1 by comparing the nearest neighbors with bootstrap analysis (bootstrap NJ tree) using 1000 bootstrap tests (1000 alternative trees). Then the phylogenetic tree was edited by the program Mega v. 6.00. Results. Based on the results of comparative, phylogenetic and phenotypic analysis, the studied Antarctic strains 180n1, 181n2, 188n2, 190n2 were identified as Pseudomonas fluorescens, Microbacterium foliorum, Sporosarcina aquimarina and Rothia sp., respectively. The coefficient of similarity of 16S rRNA genes of strain 180n1 with such a closely related species from the database P. fluorescens NBRC 14160 was 99.5%; 181n2 with M. foliorum P 333/02 – 99.4%; 188n2 with S. aquimarina SW28 – 99.7%. These strains form common clusters with closely related species on phylogenetic dendrograms. The strain 190n2 can be considered as Rothia sp., since has the remote position from closely related strains in the cluster Rothia and a low percentage of similarity (97.3%) with the species Rothia endophytica YIM 67072. These strains belong to the phyla: Firmicutes, Actinobacteria, Proteobacteria. Conclusions. Phylogenetic and phenotypic analyzes allowed determining the taxonomic position of isolated aerobic chemoorganotrophic microbial strains of the Antarctic. Nucleotide sequences of 16S rRNA genes are deposited in the International GenBank database under numbers HG518622, HG518623, HG518625, HG518626.