Habitat distribution of the genus Belliella in continental waters and the description of Belliella alkalica sp. nov., Belliella calami sp. nov. and Belliella filtrata sp. nov.

The genus Belliella belongs to the family Cyclobacteriaceae (order Cytophagales, phylum Bacteroidota) and harbours aerobic chemoheterotrophic bacteria. Members of this genus were isolated from various aquatic habitats, and our analysis based on global amplicon sequencing data revealed that their relative abundance can reach up to 5-10 % of the bacterioplankton in soda lakes and pans. Although a remarkable fraction of the most frequent genotypes that we identified from continental aquatic habitats is still uncultured, five new alkaliphilic Belliella strains were characterized in detail in this study, which were isolated from three different soda lakes and pans of the Carpathian Basin (Hungary). Cells of all strains were Gram-stain-negative, obligate aerobic, rod-shaped, non-motile and non-spore-forming. The isolates were oxidase- and catalase-positive, red-coloured, but did not contain flexirubin-type pigments; they formed bright red colonies that were circular, smooth and convex. Their major isoprenoid quinone was MK-7 and the predominant fatty acids were iso-C15 : 0, iso-C17 : 0 3-OH and summed feature 3 containing C16 : 1 ω6c and/or C16 : 1 ω7c. The polar lipid profiles contained phosphatidylethanolamine, an unidentified aminophospholipid, an unidentified glycolipid, and several unidentified lipids and aminolipids. Based on whole-genome sequences, the DNA G+C content was 37.0, 37.1 and 37.8 mol % for strains R4-6T, DMA-N-10aT and U6F3T, respectively. The distinction of three new species was confirmed by in silico genomic comparison. Orthologous average nucleotide identity (<85.4 %) and digital DNA-DNA hybridization values (<38.9 %) supported phenotypic, chemotaxonomic and 16S rRNA gene sequence data and, therefore, the following three novel species are proposed: Belliella alkalica sp. nov. (represented by strains R4-6T=DSM 111903T=JCM 34281T=UCCCB122T and S4-10), Belliella calami sp. nov. (DMA-N-10aT=DSM 107340T=JCM 34280T=UCCCB121T) and Belliella filtrata sp. nov. (U6F3T=DSM 111904T=JCM 34282T=UCCCB123T and U6F1). Emended descriptions of species Belliella aquatica, Belliella baltica, Belliella buryatensis, Belliella kenyensis and Belliella pelovolcani are also presented.

Szabonella alba gen. nov., sp. nov., a motile alkaliphilic bacterium of the family Rhodobacteraceae isolated from a soda lake.

A Gram-stain-negative, oxidase- and catalase-positive, rod-shaped, creamy white coloured bacterial strain, DMG-N-6T, was isolated from a water sample of Lake Ferto/Neusiedler See (Hungary). Phylogenetic analysis based on 16S rRNA gene sequences revealed that the strain forms a distinct linage within the family Rhodobacteraceae. Its closest relatives are Tabrizicola alkalilacus DJCT (96.76% similarity) and Tabrizicola piscis K13M18T (96.76%), followed by Tabrizicola sediminis DRYC-M-16T (96.69 %), Rhodobacter sediminicola JA983T (96.62 %), Tabrizicola aquatica RCRI19T (96.47 %) and Cereibacter johrii JA192T (96.18 %). The novel bacterial strain favours an alkaline environment (pH 8.0-12.0) and grows optimally at 18-28°C in the presence of 2-4 % (w/v) NaCl. Cells of DMG-N-6T were motile by a single subpolar flagellum. Bacteriochlorophyll a was not detected. The predominant respiratory quinone was ubiquinone Q-10. The major cellular fatty acid was C18:1 ω7c. The polar lipid profile comprised phosphatidylglycerol, diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylserine, phosphatidylcholine, an unidentified phospholipid and five unidentified lipids. The assembled draft genome of strain DMG-N-6T had 52 contigs with a total length of 4 219 778 bp and a G+C content of 64.3 mol%. Overall genome-related indices (ANI <77.8 %, AAI <69.0 %, dDDH <19.6 %) with respect to close relatives were all significantly below the corresponding threshold to demarcate bacterial genus and species. Strain DMG-N-6T (=DSM 108208T=NCAIM B.02645T) is strongly different from its closest relatives and is suggested as the type strain of a novel species of a new genus in the family Rhodobacteraceae, for which the name Szabonella alba gen. nov., sp. nov. is proposed.

Microbiological investigations of two thermal baths in Budapest, Hungary. Report: effect of bathing and pool operation type on water quality.

In Hungary, which is famous for its thermal baths, according to the regulations, waters are investigated in hygienic aspects with standard cultivation methods. In the present study, two thermal baths were investigated (the well and three different pool waters in both) using cultivation methods, taxon-specific polymerase chain reactions (PCRs), multiplex PCRs and next-generation amplicon sequencing. Mainly members of the natural microbial community of the well waters and bacteria originating from the environment were detected but several opportunistic pathogenic taxa, e.g., Pseudomonas aeruginosa, P. stutzeri, Acinetobacter johnsoni, Acinetobacter baumanni, Moraxella osloensis, Microbacterium paraoxydans, Legionella spp., Stenotrophomonas maltophilia and Staphylococcus aureus were revealed by the applied methods. Pools with charging-unloading operation had higher microscopic cell counts, colony-forming unit (CFU) counts, number of cocci, P. aeruginosa and S. aureus compared to the recirculation systems. Bacteria originating from human sources (e.g., skin) were identified in the pool waters with less than 1% relative abundance, and their presence was sporadic in the pools. Comparing the microbiological quality of the pools based on the first sampling time and the following four months' period it was revealed that recirculation operation type has better water quality than the charging-unloading pool operation from a hygienic point of view.

Prokaryotic community composition in a great shallow soda lake covered by large reed stands (Neusiedler See/Lake Ferto) as revealed by cultivation- and DNA-based analyses.

Little is known about the detailed community composition of heterotrophic bacterioplankton in macrophyte-dominated littoral systems, where a considerable amount of dissolved organic carbon originates from aquatic macrophytes instead of phytoplankton. The aim of the present study was to reveal the effect of macrophytes on the microbial community and to elucidate their role in a macrophyte-dominated shallow soda lake, which can be characterised by a mosaic of open waters and reed marsh. Therefore, 16S rRNA gene amplicon sequencing, the most probable number method, cultivation of bacterial strains, EcoPlate and cultivation-based substrate utilisation techniques were applied. Differences in the structures of microbial communities were detected between the water and the sediment samples and between vegetated and unvegetated water samples. Planktonic bacterial communities of an inner pond and a reed-covered area showed significant similarities to each other. Woesearchaeia was the dominant archaeal taxon in the water samples, while Bathyarchaeia, 'Marine Benthic Group D' and 'DHVEG-1' were abundant in the sediment samples. The most probable number of heterotrophic bacteria was lower in the open water than in the reed-associated areas. The vast majority (83%) of the isolated bacterial strains from the water samples of the reed-covered area were able to grow on a medium containing reed extract as the sole source of carbon.

Phragmitibacter flavus gen. nov., sp. nov. a new member of the family Verrucomicrobiaceae.

The Gram-stain-negative, aerobic, non-motile, oxidase- and catalase-positive, rod-shaped yellow-coloured bacterial strain MG-N-17T was isolated from a water sample of Lake Ferto/Neusiedler See (Hungary). Results of phylogenetic analysis based on the 16S rRNA gene sequence revealed that the strain forms a distinct linage within the family Verrucomicrobiaceae of the phylum Verrucomicrobia, and its closest relatives are Verrucomicrobium spinosum DSM 4136T (94.38 %) and Roseimicrobium gellanilyticum DC2a-G7T (91.55 %). The novel bacterial strain prefers a weak alkaline environment and grows optimally between 22-28 °C in the absence of NaCl. The major isoprenoid quinones are MK-10, MK-11, MK-12 and MK-9. The major cellular fatty acids are anteiso-C15 : 0, C16 : 0, C16 : 1ω5c and iso-C14 : 0. The polar lipid profile contains phosphatidylglycerol, diphosphatidylglycerol, phosphatidylethanolamine, two unidentified phospholipids and four unidentified glycolipids. The assembled draft genome of strain MG-N-17T had 44 contigs with an N50 value 348255 nt, 56.5× genome coverage, total length of 5 910 933 bp and G+C content of 56.9 mol%. Strain MG-N-17T (=DSM 106674T=NCAIM B.02643T) is proposed as the type strain of a new genus and species in the family Verrucomicrobiaceae, for which the name Phragmitibacter flavus gen. nov., sp. nov. is proposed.

Fertoeibacter niger gen. nov., sp. nov. a novel alkaliphilic bacterium of the family Rhodobacteraceae.

Three Gram-stain-negative, non-motile, oxidase- and catalase-positive, rod-shaped, black, facultative phototrophic bacterial strains, RG-N-1aT, DMA-N-7a and RA-N-9 were isolated from the water sample from Lake Ferto/Neusiedler See (Hungary). Phylogenetic analysis based on the 16S rRNA gene sequences revealed that the strains form a distinct linage within the family Rhodobacteraceae and their closest relatives are Tabrizicola piscis K13M18T (96.32%) followed by Cypionkella psychrotolerans PAMC 27389T (96.25%). The novel bacterial strains prefer alkaline environments and grow optimally at 23-33 °C in the presence of NaCl (1-2 w/v%). Bacteriochlorophyll a was detected. Cells contained exclusively ubiquinone Q-10. The major cellular fatty acids were C18 : 1ω7c, C19 : 1iso ω5c, C18 : 0 3-OH and C18 : 1ω7c 11-methyl. The polar lipid profile contains diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylcholine, an unidentified phospholipid and four unidentified lipids. The assembled draft genome of RG-N-1aT had 33 contigs with N50 values 315 027 nt, 96× genome coverage, total length of 4 326 551 bp and a DNA G+C content of 64.9%. Genome-based calculations (genome-to-genome distance and DNA G+C percentage) and pairwise amino acid identity (AAI <73.5%) indicate that RG-N-1aT represents a novel genus. RG-N-1aT (=DSM 108317T=NCAIM B.02647T) is suggested as the type strain of a novel genus and species in the family Rhodobacteraceae, for which the name Fertoeibacter niger gen. nov., sp. nov. is proposed.

Gellertiella hungarica gen. nov., sp. nov., a novel bacterium of the family Rhizobiaceae isolated from a spa in Budapest.

A novel alphaproteobacterium, strain RAM11T, belonging to the family Rhizobiaceae was isolated from the pool water of a thermal bath in Budapest, Hungary. Based on the 16S rRNA gene sequence strain RAM11T shows the highest sequence similarity values to Ensifer adhaerens Casida A (97.44 %), to Ensifer (syn. Sinorhizobium) americanus CFNEI 156T (96.87 %) and to Rhizobium azooxidifex Po 20/26T (96.76 %). The new bacterium is strictly aerobic, its optimum growth occurs at 20-37 °C, between pH 7 and 9 and without NaCl. It is motile due to a single polar flagellum, capable of budding and forms rosettes in liquid culture. The major isoprenoid quinone of strain RAM11T is Q-10, the major cellular fatty acids are C18 : 1ω7c and 11-MeC18 : 1ω7c. The polar lipid profile contains phosphatidylglycerol, phosphatidylethanolamine, phosphatidylmonomethylethanolamine, phosphatidylcholine, an unidentified aminolipid and an unidentified phospholipid. The G+C content of DNA of the type strain is 62.9 mol%. Strain RAM11T (=DSM 29853T=NCAIM B.02618T) is proposed as type strain of a new genus and species with the proposed name Gellertiella hungarica gen. nov., sp. nov.

Microbiological investigations on the water of a thermal bath at Budapest.

Thermal baths are unique aquatic environments combining a wide variety of natural and anthropogenic ecological factors, which also appear in their microbiological state. There is limited information on the microbiology of thermal baths in their complexity, tracking community shifts from the thermal wells to the pools. In the present study, the natural microbial community of well and pool waters in Gellért bath was studied in detail by cultivation-based techniques. To isolate bacteria, 10% R2A and minimal synthetic media (with "bath water") with agar-agar and gellan gum were used after prolonged incubation time; moreover, polyurethane blocks covered with media were also applied. Strains were identified by sequencing their 16S rRNA gene after grouping them by amplified rDNA restriction analysis. From each sample, the dominance of Alphaproteobacteria was characteristic though their diversity differed among samples. Members of Actinobacteria, Firmicutes, Beta- and Gamma-proteobacteria, Deinococcus-Thermus, and Bacteroidetes were also identified. Representatives of Deinococcus-Thermus phylum appeared only in the pool water. The largest groups in the pool water belonged to the Tistrella and Chelatococcus genera. The most dominant member in the well water was a new taxon, its similarity to Hartmannibacter diazotrophicus as closest relative was 93.93%.