Ulrike Koll’s research while affiliated with University of Innsbruck and other places

We present two strains affiliated with the GKS98 cluster. This phylogenetically defined cluster is representing abundant, mainly uncultured freshwater bacteria, which were observed by many cultivation-independent studies on the diversity of bacteria in various freshwater lakes and streams. Bacteria affiliated with the GKS98 cluster were detected by cultivation-independent methods in freshwater systems located in Europe, Asia, Africa and the Americas. The two strains, LF4-65 T (=CCUG 56422 T =DSM 107630 T ) and MWH-P2sevCIIIb T (=CCUG 56420 T =DSM 107629 T ), are aerobic chemoorganotrophs, both with genome sizes of 3.2 Mbp and G+C values of 52.4 and 51.0 mol%, respectively. Phylogenomic analyses based on concatenated amino acid sequences of 120 proteins suggest an affiliation of the two strains with the family Alcaligenaceae and revealed Orrella amnicola and Orrella marina (= Algicoccus marinus ) as being the closest related, previously described species. However, the calculated phylogenomic trees clearly suggest that the current genus Orrella represents a polyphyletic taxon. Based on the branching order in the phylogenomic trees, as well as the revealed phylogenetic distances and chemotaxonomic traits, we propose to establish the new genus Zwartia gen. nov. and the new species Z. hollandica sp. nov. to harbour strain LF4-65 T and the new genus Jezberella gen. nov. and the new species J. montanilacus sp. nov. to harbour strain MWH-P2sevCIIIb T . Furthermore, we propose the reclassification of the species Orrella amnicola in the new genus Sheuella gen. nov. The new genera Zwartia , Jezberella and Sheuella together represent taxonomically the GKS98 cluster.

Two bacterial strains, 9H-EGSE T and 15D-MOB T , were isolated from small freshwater habitats located near Salzburg, Austria. They showed the highest 16S rRNA sequence similarities of 100% and 99.9%, respectively, with type strains of species of the genus Aquirufa ( Bacteroidota ). Genome-based phylogenetic reconstructions with 119 amino acid sequences assigned the new taxa to the two distinct branches of the genus Aquirufa . Whole-genome average nucleotide identities were calculated with all possible pairs belonging to the genus. Values between 75.4% and 88.6% revealed that the two new strains represent each a new species. Like all, so far described members of the genus, they grew aerobically and chemoorganotrophically, were rod-shaped, red-pigmented, and motile by gliding, and showed genome sizes of about 3 Mbp and G + C values of about 40%. They could be distinguished by some phenotypic and chemotaxonomic features from their nearest related species. Until now, strain 9H-EGSE T is the only one among the Aquirufa strains which contained traces of MK8 as respiratory quinone, and strain 15D-MOB T is the only one that formed tiny orange globules in liquid medium. The genome of strain 9H-EGSE T comprised genes for the complete light-harvesting rhodopsin / retinal system, in the case of 15D-MOB T genes predicted for a nitrous oxide reductase were present. For the two new species of the genus Aquirufa , we propose to establish the names Aquirufa lenticrescens for strain 9H-EGSE T (= JCM 34077 T = CIP 111926 T ) and Aquirufa aurantiipilula for strain 15D-MOB T (= JCM 34078 T = CIP 111925 T ).

The aerobic primarily chemoorganotrophic actinobacterial strain MWH-Mo1 T was isolated from a freshwater lake and is characterized by small cell lengths of less than 1 µm, small cell volumes of 0.05–0.06 µm ³ (ultramicrobacterium), a small genome size of 1.75 Mbp and, at least for an actinobacterium, a low DNA G+C content of 54.6 mol%. Phylogenetic analyses based on concatenated amino acid sequences of 116 housekeeping genes suggested the type strain of Aurantimicrobium minutum affiliated with the family Microbacteriaceae as its closest described relative. Strain MWH-Mo1 T shares with the type strain of that species a 16S rRNA gene sequence similarity of 99.6 % but the genomes of the two strains share an average nucleotide identity of only 79.3 %. Strain MWH-Mo1 T is in many genomic, phenotypic and chemotaxonomic characteristics quite similar to the type strain of A. minutum . Previous intensive investigations revealed two unusual traits of strain MWH-Mo1 T . Although the strain is not known to be phototrophic, the metabolism is adjusted to the diurnal light cycle by up- and down-regulation of genes in light and darkness. This results in faster growth in the presence of light. Additionally, a cell size-independent protection against predation by bacterivorous flagellates, most likely mediated by a proteinaceous cell surface structure, was demonstrated. For the previously intensively investigated aerobic chemoorganotrophic actinobacterial strain MWH-Mo1 T (=CCUG 56426 T =DSM 107758 T ), the establishment of the new species Aurantimicrobium photophilum sp. nov. is proposed.

The actinobacterial strain 15G-AUS-rot T was isolated from an artificial pond located near Salzburg, Austria. The strain showed 16S rRNA gene sequence similarities of 98.7 % to Candidatus Aquiluna rubra and of 96.6 and 96.7 % to the two validly described species of the genus Rhodoluna . Phylogenetic reconstructions based on 16S rRNA gene sequences and genome-based on amino acid sequences of 118 single copy genes referred strain 15G-AUS-rot T to the family Microbacteriaceae and therein to the so-called subcluster Luna-1. The genome-based phylogenetic tree showed that the new strain represents a putative new genus. Cultures of strain 15G-AUS-rot T were light red pigmented and comprised very small, rod-shaped cells. They metabolized a broad variety of substrates. Major fatty acids (>10 %) of cells were iso-C 16 : 0 , antiso-C 15 : 0 and iso-C 14 : 0 . The major respiratory quinone was MK-11 and a minor component was MK-10. The peptidoglycan structure belonged to an unusual B type. The closed genome sequence of the strain was very small (1.4 Mbp) and had a DNA G+C content of 54.8 mol%. An interesting feature was the presence of genes putatively encoding the complete light-driven proton pumping actinorhodopsin/retinal system, which were located at three different positions of the genome. Based on the characteristics of the strain, a new genus and a new species termed Aquiluna borgnonia is proposed for strain 15G-AUS-rot T (=DSM 107803 T =JCM 32974 T ).

Two bacterial strains, 50A-KIRBA T and 50C-KIRBA T , were isolated from the same freshwater creek located near Salzburg, Austria. They showed 16S rRNA gene sequence similarities to Aquirufa nivalisilvae of 100 and 99.9 %, respectively. A genome-based phylogenetic reconstruction with amino acid sequences of 119 single-copy genes suggested that the new strains represent two new species of the genus Aquirufa . Pairwise calculated whole-genome average nucleotide identity (gANI) values ranging from 85.4 to 87.5 % confirmed this conclusion. Phenotypic, chemotaxonomic and genomic traits were investigated. Like strains of other Aquirufa species, 50A-KIRBA T and 50C-KIRBA T grew aerobically and chemoorganotrophically, were rod-shaped, red-pigmented and motile, most likely by gliding. They could be distinguished by slight differences in the chemotaxonomic features. We propose to establish for strain 50A-KIRBA T (=CIP 111735 T =LMG 31080 T ) as type strain the name Aquirufa ecclesiirivi and for strain 50C-KIRBA T (=CIP 111736 T =LMG 31501 T ) as type strain the name Aquirufa beregesia . Furthermore, the relationship between the type strains of Aquirufa nivalisilvae (59G-WUEMPEL T ) and Allopseudarcicella aquatilis (HME7025 T ) was investigated. Results of polyphasic analyses, especially a gANI value of 97.6 %, as well as the genome-based phylogenetic reconstruction, suggested that Allopseudarcicella aquatilis is a heterotypic synonym of Aquirufa nivalisilvae . According to rule 24b of the International Code of Nomenclature of Prokaryotes we propose to classify strain HME7025 as Aquirufa nivalisilvae and provide an emended description for the latter.

The bacterial strain 53C-WASEF was isolated from a small freshwater ditch located in Eugendorf, Austria. Phylogenetic reconstructions with 16S rRNA gene sequences and genome based, with amino acid sequences obtained from 105 single copy genes, suggested that the strain represents a new genus and a new species within the family Opitutaceae, which belongs to the class Opitutae of the phylum Verrucomicrobia. Comparisons of the 16S rRNA gene sequence of strain 53C-WASEF with those of related type strains revealed a highest sequence similarity of 93.5 % to Nibricoccus aquaticus and of 92.9 % to Geminisphaera colitermitum. Interestingly, phylogentic trees indicated the latter as being the closest known relative of the new strain. Phenotypic, chemotaxonomic and genomic traits were investigated. Cells were observed to be small, spherical, motile and unpigmented, and grew chemoorganotrophically and aerobically. The respiratory quinone was MK-7, the predominant fatty acids were anteiso-C15 : 0, C16 : 1ω5c and C16 : 0. The identified polar lipids were phosphatidylethanolamine, phosphatidylglycerol and diphosphatidylglycerol. Genome sequencing revealed genes putatively encoding for flagella synthesis and cellulose degradation. The genome size was 4.1 Mbp and the G+C content 60.6 mol%. For the new genus and the new species, we propose the name Rariglobus hedericola gen. nov., sp. nov. (=CIP 111665T=DSM 109123T).

Strain 33A1-SZDPT was isolated from a small creek located in Puch, Austria. Strain SP-Ram-0.45-NSY-1T was obtained from a small pond located in Schönramer Moor, Germany. 16S rRNA gene sequence similarities between the type strain of Silvanigrella aquatica, currently the only member of the family Silvanigrellaceae, and strains 33A1-SZDPT and SP-Ram-0.45-NSY-1T of 94.1 and 99.1 %, respectively, suggested affiliation of the two strains with this family. Phylogenetic reconstructions with 16S rRNA gene sequences and phylogenomic analyses with amino acid sequences obtained from 103 single-copy genes suggested that the strains represent a new genus and a new species in the case of strain 33A1-SZDPT (=JCM 32978T=DSM 107810T), and a new species within the genus Silvanigrella in the case of strain SP-Ram-0.45-NSY-1T (=JCM 32975T=DSM 107809T). Cells of strain 33A1-SZDPT were motile, pleomorphic, purple-pigmented on agar plates, putatively due to violacein, and showed variable pigmentation in liquid media. They grew chemoorganotrophically and aerobically and tolerated salt concentrations up to 1.2 % NaCl (v/w). The genome size of strain 33A1-SZDPT was 3.4 Mbp and the G+C content was 32.2 mol%. For this new genus and new species, we propose the name Fluviispira multicolorata gen. nov., sp. nov. Cells of strain SP-Ram-0.45-NSY-1T were motile, pleomorphic, red-pigmented and grew chemoorganotrophically and aerobically. They tolerated salt concentrations up to 1.1 % NaCl (v/w). The genome size of strain SP-Ram-0.45-NSY-1T was 3.9 Mbp and the G+C content 29.3 mol%. For the new species within the genus Silvanigrella we propose the name Silvanigrella paludirubra sp. nov.

Three actinobacterial strains, 27D-LEPIT, 1B-Mac and 36A-HELLB, were isolated from small standing and running freshwater habitats located in Salzburg, Austria. Phylogenetic reconstructions based on 16S rRNA gene sequences and genome based on concatenated amino acid sequences of 119 single-copy genes referred the three strains within the family Microbacteriaceae to the genus Rhodoluna. The strains showed 100 % 16S rRNA gene sequence similarities to the previously described Candidatus Rhodoluna limnophila. Cells were very small, approximately 0.5×0.3 µm, and showed a red pigmentation in liquid nutrient broth-soyotone-yeast extract medium as well as on agar plates. The strains assimilated a broad variety of substrates; however, the patterns differed a great deal between the three investigated strains. For strain 27D-LEPIT, the major fatty acids were iso-C14 : 0 and anteiso-C15 : 0; the identified polar lipids were phosphatidylglycerol and diphosphatidylglycerol; the major respiratory quinone was MK-11; and analysis of the peptidoglycan structure indicated the presence of type B2β (typeB10). The genomic DNA of the three strains was very small (1.4 Mbp) and had a DNA G+C content of 54 mol%. The investigated traits suggested that strains 36A-HELLB (=DSM 107805=JCM 32927), 1B-Mac (=DSM 107802=JCM 32925) and 27D-LEPIT (=JCM 32926T =DSM 107804T) represent a new species for which the name Rhodolunalimnophila sp. nov. is proposed, with strain 27D-LEPIT as the type strain.

Three bacterial strains, 30S-ANTBAC, 103A-SOEBACH and 59G- WUEMPEL, were isolated from two small freshwater creeks and an intermittent pond near Salzburg, Austria. Phylogenetic reconstructions with 16S rRNA gene sequences and, genome based, with amino acid sequences obtained from 119 single copy genes showed that the three strains represent a new genus of the family Cytophagaceae within a clade formed by the genera Pseudarcicella, Arcicella and Flectobacillus. blast searches suggested that the new genus comprises widespread freshwater bacteria. Phenotypic, chemotaxonomic and genomic traits were investigated. Cells were rod shaped and were able to glide on soft agar. All strains grew chemoorganotrophically and aerobically, were able to assimilate pectin and showed an intense red pigmentation putatively due to various carotenoids. Two strains possessed genes putatively encoding proteorhodopsin and retinal biosynthesis. Genome sequencing revealed genome sizes between 2.5 and 3.1 Mbp and G+C contents between 38.0 and 42.7 mol%. For the new genus we propose the name Aquirufa gen. nov. Pairwise-determined whole-genome average nucleotide identity values suggested that the three strains represent two new species within the new genus for which we propose the names Aquirufa antheringensis sp. nov. for strain 30S-ANTBACT (=JCM 32977T =LMG 31079T=DSM 108553T) as type species of the genus, to which also belongs strain 103A-SOEBACH (=DSM 108555=LMG 31082) and Aquirufa nivalisilvae sp. nov. for strain 59G-WUEMPELT (=LMG 31081T =DSM 108554T).