Carnobacterium iners sp. nov., a psychrophilic, lactic acid-producing bacterium from the littoral zone of an Antarctic pond.

Two lactic acid-producing, Gram-stain-positive rods were isolated from a microbial mat actively growing in the littoral zone of an Antarctic lake (Forlidas Pond) in the Pensacola mountains and studied using a polyphasic taxonomic approach. The isolates were examined by phylogenetic analysis of the 16S rRNA gene, multilocus sequence analysis of pheS, rpoA and atpA, and biochemical and genotypic characteristics. One strain, designated LMG 26641, belonged to Carnobacterium alterfunditum and the other strain, designated LMG 26642(T), could be assigned to a novel species, with Carnobacterium funditum DSM 5970(T) as its closest phylogenetic neighbour (99.2 % 16S rRNA gene sequence similarity). Carnobacterium iners sp. nov. could be distinguished biochemically from other members of the genus Carnobacterium by the lack of acid production from carbohydrates. DNA-DNA relatedness confirmed that strain LMG 26642(T) represented a novel species, for which we propose the name Carnobacterium iners sp. nov. (type strain is LMG 26642(T)  = CCUG 62000(T)).

Culturable diversity of heterotrophic bacteria in Forlidas Pond (Pensacola Mountains) and Lundström Lake (Shackleton Range), Antarctica.

Cultivation techniques were used to study the heterotrophic bacterial diversity in two microbial mat samples originating from the littoral zone of two continental Antarctic lakes (Forlidas Pond and Lundström Lake) in the Dufek Massif (within the Pensacola Mountains group of the Transantarctic Mountains) and Shackleton Range, respectively. Nearly 800 isolates were picked after incubation on several growth media at different temperatures. They were grouped using a whole-genome fingerprinting technique, repetitive element palindromic PCR and partial 16S rRNA gene sequencing. Phylogenetic analysis of the complete 16S rRNA gene sequences of 82 representatives showed that the isolates belonged to four major phylogenetic groups: Actinobacteria, Bacteroidetes, Proteobacteria and Firmicutes. A relatively large difference between the samples was apparent. Forlidas Pond is a completely frozen water body underlain by hypersaline brine, with summer thaw forming a slightly saline littoral moat. This was reflected in the bacterial diversity with a dominance of isolates belonging to Firmicutes, whereas isolates from the freshwater Lundström Lake revealed a dominance of Actinobacteria. A total of 42 different genera were recovered, including first records from Antarctica for Albidiferax, Bosea, Curvibacter, Luteimonas, Ornithinibacillus, Pseudoxanthomonas, Sphingopyxis and Spirosoma. Additionally, a considerable number of potential new species and new genera were recovered distributed over different phylogenetic groups. For several species where previously only the type strain was available in cultivation, we report additional strains. Comparison with public databases showed that overall, 72% of the phylotypes are cosmopolitan whereas 23% are currently only known from Antarctica. However, for the Bacteroidetes, the majority of the phylotypes recovered are at present known only from Antarctica and many of these represent previously unknown species.

Bacterial community composition in relation to bedrock type and macrobiota in soils from the Sør Rondane Mountains, East Antarctica.

Antarctic soils are known to be oligotrophic and of having low buffering capacities. It is expected that this is particularly the case for inland high-altitude regions. We hypothesized that the bedrock type and the presence of macrobiota in these soils enforce a high selective pressure on their bacterial communities. To test this, we analyzed the bacterial community structure in 52 soil samples from the western Sør Rondane Mountains (Dronning Maud Land, East Antarctica), using the Illumina MiSeq platform in combination with ARISA fingerprinting. The samples were taken along broad environmental gradients in an area covering nearly 1000 km(2) Ordination and variation partitioning analyses revealed that the total organic carbon content was the most significant variable in structuring the bacterial communities, followed by pH, electric conductivity, bedrock type and the moisture content, while spatial distance was of relatively minor importance. Acidobacteria (Chloracidobacteria) and Actinobacteria (Actinomycetales) dominated gneiss derived mineral soil samples, while Proteobacteria (Sphingomonadaceae), Cyanobacteria, Armatimonadetes and candidate division FBP-dominated soil samples with a high total organic carbon content that were mainly situated on granite derived bedrock.

Bacterial diversity assessment in Antarctic terrestrial and aquatic microbial mats: a comparison between bidirectional pyrosequencing and cultivation.

The application of high-throughput sequencing of the 16S rRNA gene has increased the size of microbial diversity datasets by several orders of magnitude, providing improved access to the rare biosphere compared with cultivation-based approaches and more established cultivation-independent techniques. By contrast, cultivation-based approaches allow the retrieval of both common and uncommon bacteria that can grow in the conditions used and provide access to strains for biotechnological applications. We performed bidirectional pyrosequencing of the bacterial 16S rRNA gene diversity in two terrestrial and seven aquatic Antarctic microbial mat samples previously studied by heterotrophic cultivation. While, not unexpectedly, 77.5% of genera recovered by pyrosequencing were not among the isolates, 25.6% of the genera picked up by cultivation were not detected by pyrosequencing. To allow comparison between both techniques, we focused on the five phyla (Proteobacteria, Actinobacteria, Bacteroidetes, Firmicutes and Deinococcus-Thermus) recovered by heterotrophic cultivation. Four of these phyla were among the most abundantly recovered by pyrosequencing. Strikingly, there was relatively little overlap between cultivation and the forward and reverse pyrosequencing-based datasets at the genus (17.1-22.2%) and OTU (3.5-3.6%) level (defined on a 97% similarity cut-off level). Comparison of the V1-V2 and V3-V2 datasets of the 16S rRNA gene revealed remarkable differences in number of OTUs and genera recovered. The forward dataset missed 33% of the genera from the reverse dataset despite comprising 50% more OTUs, while the reverse dataset did not contain 40% of the genera of the forward dataset. Similar observations were evident when comparing the forward and reverse cultivation datasets. Our results indicate that the region under consideration can have a large impact on perceived diversity, and should be considered when comparing different datasets. Finally, a high number of OTUs could not be classified using the RDP reference database, suggesting the presence of a large amount of novel diversity.

The gyrB gene is a useful phylogenetic marker for exploring the diversity of Flavobacterium strains isolated from terrestrial and aquatic habitats in Antarctica.

Within the phylum Bacteroidetes, the gyrB gene, encoding for the B subunit of the DNA gyrase, has been used as a phylogenetic marker for several genera closely related to Flavobacterium. The phylogenies of the complete 16S rRNA gene and the gyrB gene were compared for 33 Antarctic Flavobacterium isolates and 23 type strains from closely related Flavobacterium species. gyrB gene sequences provided a higher discriminatory power to distinguish between different Flavobacterium groups than 16S rRNA gene sequences. The gyrB gene is therefore a promising molecular marker for elucidating the phylogenetic relationships among Flavobacterium species and should be evaluated for all the other type strains of described Flavobacterium species. Combining the phylogeny of both genes, the new Antarctic Flavobacterium strains constitute 15 Flavobacterium groups, including at least 13 potentially new species together with one group of isolates probably belonging to the species Flavobacterium micromati and one group close to Flavobacterium gelidilacus.

Culturable bacterial diversity at the Princess Elisabeth Station (Utsteinen, Sør Rondane Mountains, East Antarctica) harbours many new taxa.

We studied the culturable heterotrophic bacterial diversity present at the site of the new Princess Elisabeth Station at Utsteinen (Dronning Maud Land, East Antarctica) before construction. About 800 isolates were picked from two terrestrial microbial mat samples after incubation on several growth media at different temperatures. They were grouped using rep-PCR fingerprinting and partial 16S rRNA gene sequencing. Phylogenetic analysis of the complete 16S rRNA gene sequences of 93 representatives showed that the isolates belonged to five major phyla: Actinobacteria, Bacteroidetes, Proteobacteria, Firmicutes and Deinococcus-Thermus. Isolates related to the genus Arthrobacter were the most prevalent whereas the genera Hymenobacter, Deinococcus, Cryobacterium and Sphingomonas were also recovered in high numbers in both samples. A total of 35 different genera were found, the majority of which has previously been reported from Antarctica. For the genera Aeromicrobium, Aurantimonas, Rothia, Subtercola, Tessaracoccus and Xylophilus, this is the first report in Antarctica. In addition, numerous potential new species and new genera were recovered; many of them currently restricted to Antarctica, particularly in the phyla Bacteroidetes and Deinococcus-Thermus.