Priority effects alter microbiome composition and increase abundance of probiotic taxa in treefrog tadpoles.

Host-associated microbial communities, like other ecological communities, may be impacted by the colonization order of taxa through priority effects. Developing embryos and their associated microbiomes are subject to stochasticity during colonization by bacteria. For amphibian embryos, often developing externally in bacteria-rich environments, this stochasticity may be particularly impactful. For example, the amphibian microbiome can mitigate lethal outcomes from disease for their hosts; however, this may depend on microbiome composition. Here, we examined the assembly of the bacterial community in spring peeper (Pseudacris crucifer) embryos and tadpoles. First, we reared embryos from identified mating pairs in either lab or field environments to examine the relative impact of environment and parentage on embryo and tadpole bacterial communities. Second, we experimentally inoculated embryos to determine if priority effects (i) could be used to increase the relative abundance of Janthinobacterium lividum, an amphibian-associated bacteria capable of preventing fungal infection, and (ii) would lead to observed differences in the relative abundances of two closely related bacteria from the genus Pseudomonas. Using 16S rRNA gene amplicon sequencing, we observed differences in community composition based on rearing location and parentage in embryos and tadpoles. In the inoculation experiment, we found that priority inoculation could increase the relative abundance of J. lividum, but did not find that either Pseudomonas isolate was able to prevent colonization by the other when given priority. These results highlight the importance of environmental source pools and parentage in determining microbiome composition, while also providing novel methods for the administration of a known amphibian probiotic. IMPORTANCE: Harnessing the functions of host-associated bacteria is a promising mechanism for managing disease outcomes across different host species. In the case of amphibians, certain frog-associated bacteria can mitigate lethal outcomes of infection by the fungal pathogen Batrachochytrium dendrobatidis. Successful probiotic applications require knowledge of community assembly and an understanding of the ecological mechanisms that structure these symbiotic bacterial communities. In our study, we show the importance of environment and parentage in determining bacterial community composition and that community composition can be influenced by priority effects. Further, we provide support for the use of bacterial priority effects as a mechanism to increase the relative abundance of target probiotic taxa in a developing host. While our results show that priority effects are not universally effective across all host-associated bacteria, our ability to increase the relative abundance of specific probiotic taxa may enhance conservation strategies that rely on captive rearing of endangered vertebrates.

Plant flavones enrich rhizosphere Oxalobacteraceae to improve maize performance under nitrogen deprivation.

Beneficial interactions between plant roots and rhizosphere microorganisms are pivotal for plant fitness. Nevertheless, the molecular mechanisms controlling the feedback between root architecture and microbial community structure remain elusive in maize. Here, we demonstrate that transcriptomic gradients along the longitudinal root axis associate with specific shifts in rhizosphere microbial diversity. Moreover, we have established that root-derived flavones predominantly promote the enrichment of bacteria of the taxa Oxalobacteraceae in the rhizosphere, which in turn promote maize growth and nitrogen acquisition. Genetic experiments demonstrate that LRT1-mediated lateral root development coordinates the interactions of the root system with flavone-dependent Oxalobacteraceae under nitrogen deprivation. In summary, these experiments reveal the genetic basis of the reciprocal interactions between root architecture and the composition and diversity of specific microbial taxa in the rhizosphere resulting in improved plant performance. These findings may open new avenues towards the breeding of high-yielding and nutrient-efficient crops by exploiting their interaction with beneficial soil microorganisms.

Repeated Exposure of Aspergillus niger Spores to the Antifungal Bacterium Collimonas fungivorans Ter331 Selects for Delayed Spore Germination.

The bacterial strain Collimonas fungivorans Ter331 (CfTer331) inhibits mycelial growth and spore germination in Aspergillus niger N402 (AnN402). The mechanisms underlying this antagonistic bacterial-fungal interaction have been extensively studied, but knowledge on the long-term outcome of this interaction is currently lacking. Here, we used experimental evolution to explore the dynamics of fungal adaptation to recurrent exposure to CfTer331. Specifically, five single-spore isolates (SSIs) of AnN402 were evolved under three selection scenarios in liquid culture, i.e., (i) in the presence of CfTer331 for 80 growth cycles, (ii) in the absence of the bacterium for 80 cycles, and (iii) in the presence of CfTer331 for 40 cycles and then in its absence for 40 cycles. The evolved SSI lineages were then evaluated for phenotypic changes from the founder fungal strain, such as germinability with or without CfTer331. The analysis showed that recurrent exposure to CfTer331 selected for fungal lineages with reduced germinability and slower germination, even in the absence of CfTer331. In contrast, when AnN402 evolved in the absence of the bacteria, lineages with increased germinability and faster germination were favored. SSIs that were first evolved in the presence of CfTer331 and then in its absence showed intermediate phenotypes but overall were more similar to SSIs that evolved in the absence of CfTer331 for 80 cycles. This suggests that traits acquired from exposure to CfTer331 were reversible upon removal of the selection pressure. Overall, our study provides insights into the effects on fungi from the long-term coculture with bacteria. IMPORTANCE The use of antagonistic bacteria for managing fungal diseases is becoming increasingly popular, and thus there is a need to understand the implications of their long-term use against fungi. Most efforts have so far focused on characterizing the antifungal properties and mode of action of the bacterial antagonists, but the possible outcomes of the persisting interaction between antagonistic bacteria and fungi are not well understood. In this study, we used experimental evolution in order to explore the evolutionary aspects of an antagonistic bacterial-fungal interaction, using the antifungal bacterium Collimonas fungivorans and the fungus Aspergillus niger as a model system. We show that evolution in the presence or absence of the bacteria selects for fungal lineages with opposing and conditionally beneficial traits, such as slow and fast spore germination, respectively. Overall, our studies reveal that fungal responses to biotic factors related to antagonism could be to some extent predictable and reversible.

Cooperative Interaction of Janthinobacterium sp. SLB01 and Flavobacterium sp. SLB02 in the Diseased Sponge Lubomirskia baicalensis.

Endemic freshwater sponges (demosponges, Lubomirskiidae) dominate in Lake Baikal, Central Siberia, Russia. These sponges are multicellular filter-feeding animals that represent a complex consortium of many species of eukaryotes and prokaryotes. In recent years, mass disease and death of Lubomirskia baicalensis has been a significant problem in Lake Baikal. The etiology and ecology of these events remain unknown. Bacteria from the families Flavobacteriaceae and Oxalobacteraceae dominate the microbiomes of diseased sponges. Both species are opportunistic pathogens common in freshwater ecosystems. The aim of our study was to analyze the genomes of strains Janthinobacterium sp. SLB01 and Flavobacterium sp. SLB02, isolated from diseased sponges to identify the reasons for their joint dominance. Janthinobacterium sp. SLB01 attacks other cells using a type VI secretion system and suppresses gram-positive bacteria with violacein, and regulates its own activity via quorum sensing. It produces floc and strong biofilm by exopolysaccharide biosynthesis and PEP-CTERM/XrtA protein expression. Flavobacterium sp. SLB02 utilizes the fragments of cell walls produced by polysaccharides. These two strains have a marked difference in carbohydrate acquisition. We described a possible means of joint occupation of the ecological niche in the freshwater sponge microbial community. This study expands the understanding of the symbiotic relationship of microorganisms with freshwater Baikal sponges.

Description of Massilia rubra sp. nov., Massilia aquatica sp. nov., Massilia mucilaginosa sp. nov., Massilia frigida sp. nov., and one Massilia genomospecies isolated from Antarctic streams, lakes and regoliths.

Bacteria of the genus Massilia often colonize extreme ecosystems, however, a detailed study of the massilias from the Antarctic environment has not yet been performed. Here, sixty-four Gram-stain-negative, aerobic, motile rods isolated from different environmental samples on James Ross Island (Antarctica) were subjected to a polyphasic taxonomic study. The psychrophilic isolates exhibited slowly growing, moderately slimy colonies revealing bold pink-red pigmentation on R2A agar. The set of strains exhibited the highest 16S rRNA gene sequence similarities (99.5-99.9%) to Massilia violaceinigra B2T and Massilia atriviolacea SODT and formed several phylogenetic groups based on the analysis of gyrB and lepA genes. Phenotypic characteristics allowed four of them to be distinguished from each other and from their closest relatives. Compared to the nearest phylogenetic neighbours the set of six genome-sequenced representatives exhibited considerable phylogenetic distance at the whole-genome level. Bioinformatic analysis of the genomic sequences revealed a high number of putative genes involved in oxidative stress response, heavy-metal resistance, bacteriocin production, the presence of putative genes involved in nitrogen metabolism and auxin biosynthesis. The identification of putative genes encoding aromatic dioxygenases suggests the biotechnology potential of the strains. Based on these results four novel species and one genomospecies of the genus Massilia are described and named Massilia rubra sp. nov. (P3094T=CCM 8692T=LMG 31213T), Massilia aquatica sp. nov. (P3165T=CCM 8693T=LMG 31211T), Massilia mucilaginosa sp. nov. (P5902T=CCM 8733T=LMG 31210T), and Massilia frigida sp. nov. (P5534T=CCM 8695T=LMG 31212T).

Microbial Species Coexistence Depends on the Host Environment.

Organisms and their resident microbial communities form a complex and mostly stable ecosystem. It is known that the specific composition and abundance of certain bacterial species affect host health and fitness, but the processes that lead to these microbial patterns are unknown. We investigate this by deconstructing the simple microbiome of the freshwater polyp Hydra We contrast the performance of its two main bacterial associates, Curvibacter and Duganella, on germfree hosts with two in vitro environments over time. We show that interactions within the microbiome but also the host environment lead to the observed species frequencies and abundances. More specifically, we find that both microbial species can only stably coexist in the host environment, whereas Duganella outcompetes Curvibacter in both in vitro environments irrespective of initial starting frequencies. While Duganella seems to benefit through secretions of Curvibacter, its competitive effect on Curvibacter depends upon direct contact. The competition might potentially be mitigated through the spatial distribution of the two microbial species on the host, which would explain why both species stably coexist on the host. Interestingly, the relative abundances of both species on the host do not match the relative abundances reported previously nor the overall microbiome carrying capacity as reported in this study. Both observations indicate that rare microbial community members might be relevant for achieving the native community composition and carrying capacity. Our study highlights that for dissecting microbial interactions the specific environmental conditions need to be replicated, a goal difficult to achieve with in vitro systems.IMPORTANCE This work studies microbial interactions within the microbiome of the simple cnidarian Hydra and investigates whether microbial species coexistence and community stability depend on the host environment. We find that the outcome of the interaction between the two most dominant bacterial species in Hydra's microbiome differs depending on the environment and results in a stable coexistence only in the host context. The interactive ecology between the host and the two most dominant microbes, but also the less abundant members of the microbiome, is critically important for achieving the native community composition. This indicates that the metaorganism environment needs to be taken into account when studying microbial interactions.

Glaciimonas soli sp. nov., a soil bacterium isolated from the forest of a high elevation mountain.

A Gram-negative, psychrophilic bacterium, designated strain GS1T, was isolated from a forest soil sample collected from the West Peak of Mt. Yushan, Yushan National Park, Taiwan. Cells grown in broth cultures were mostly non-motile and non-flagellated, whereas motile cells with monotrichous, subpolar flagella were also observed. The novel strain grew over a temperature range of 4-25 °C with optimum growth at 10-15 °C. It grew aerobically and was not capable of anaerobic growth by fermentation of D-glucose or other carbohydrates. Ubiquinone 8 was the predominant isoprenoid quinone. The major polar lipids comprised phosphatidylethanolamine, diphosphatidylglycerol and dimethylaminoethanol. Cellular fatty acids were dominated by C16:1ω7c (35.2%), C16:0 (19.5%), C18:1ω7c (18.8%) and C17:0ω7c cyclo (15.5%). The DNA G + C content was 49.2 mol% evaluated according to the genomic sequencing data. Strain GS1T shared more than 96.5% 16S rRNA gene sequence similarities with type strains of four Collimonas species (97.2-97.5%), three Glaciimonas species (97.3% for each of the three) and Oxalicibacterium solurbis (96.5%). Phylogenetic analysis based on 16S rRNA gene sequences revealed that strain GS1T formed a stable genus-level clade with type strains of species in the genus Glaciimonas in the family Oxalobacteraceae and GS1T was an outgroup with respect to these Glaciimonas species. Characteristically, strain GS1T could be easily distinguished from the recognised Glaciimonas species by exhibition of swimming motility with monotrichous, subpolar flagellum in some of the cells, ability to grow in NaCl at 2% but not at 3% and the distinguishable fatty acid profiles. On the basis of the polyphasic taxonomic data from this study, strain GS1T is considered to represent a novel species of the genus Glaciimonas, for which the name Glaciimonas soli sp. nov. is proposed. The type strain is GS1T (= JCM 33275T = BCRC 81091T).

Duganella rivus sp. nov., Duganella fentianensis sp. nov., Duganella qianjiadongensis sp. nov. and Massilia guangdongensis sp. nov., isolated from subtropical streams in China and reclassification of all species within genus Pseudoduganella.

Four Gram-stain-negative, catalase-positive, rod-shaped and motile strains (FT55WT, FT93WT, CY13WT and DS3T) were isolated from subtropical streams in China. Comparisons based on 16S rRNA gene sequences indicated that strains FT55WT, FT93WT and CY13WT take strain Pseudoduganella danionis E3/2T, and strain DS3T takes strain Pseudoduganella eburnea 10R5-21T as their closest neighbour, respectively. The genome sizes of strains FT55WT, FT93WT, CY13WT and DS3T were 6.15, 5.10, 5.31 and 5.72 Mbp with G+C contents of 61.7, 60.9, 60.6 and 64.0%, respectively. The reconstructed phylogenomic tree based on concatenated 92 core genes showed that strain FT55WT clusters closely with Duganella radicis KCTC 22382T and Duganella sacchari Sac-22T, strains FT93WT and CY13WT form a distinct clade with P. danionis DSM 103461T and this clade clusters with the clades of genus Duganella together, and strain DS3T forms a distinct clade with P. eburnea JCM 31587T and Pseudoduganella violaceinigra DSM 15887T and this clade clusters closely with the clades of genus Massilia, respectively. The calculated pairwise OrthoANIu values and digital DNA-DNA hybridization (DDH) values among strains FT55WT, FT93WT, CY13WT, DS3T and related strains were in the ranges of 75.6-94.2% and 20.6-56.2%, respectively. Q-8 was the sole respiratory quinone of these four strains. The major fatty acids were C16:1ω7c, C16:0 and C12:0. The polar lipids included phosphatidylglycerol, phosphatidylethanolamine and one unidentified phospholipid. Considering the similar fatty acids and polar lipids profiles of species within genus Pseudoduganella, Massilia and Duganella, there is currently no justification for assigning the species of genus Pseudoduganella into the Massilia and Duganella clades in the phylogenomic tree. It is reasonable to transfer P. violaceinigra and P. eburnea to the genus Massilia as Massilia violaceinigrum comb. nov. and Massilia eburnea comb. nov., and transfer P. danionis to the genus Duganella as Duganella danionis comb. nov. Considering phylogenomic analysis, OrthoANIu data, digital DDH data and a range of physiological and biochemical characteristics, strains FT55WT, FT93WT and CY13WT should be assigned to genus Duganella, and strain DS3T should be classified as a novel species within genus Massilia, for which the names Duganella rivus sp. nov. (type strain FT55WT = GDMCC 1.1675T = KACC 21467T), Duganella fentianensis sp. nov. (type strain FT93WT = GDMCC 1.1683T = KACC 21475T), Duganella qianjiadongensis sp. nov. (type strain CY13WT = GDMCC 1.1669T = KACC 21461T) and Massilia guangdongensis sp. nov. (type strain DS3T = GDMCC 1.1636T = KACC 21312T) are proposed.

Optimal surface estimation and thresholding of confocal microscope images of biofilms using Beer's Law.

Beer's Law explains how light attenuates into thick specimens, including thick biofilms. We use a Bayesian optimality criterion, the maximum of the posterior probability distribution, and computationally efficiently fit Beer's Law to the 3D intensity data collected from thick living biofilms by a confocal scanning laser microscope. Using this approach the top surface of the biofilm and an optimal image threshold can be estimated. Biofilm characteristics, such as bio-volumes, can be calculated from this surface. Results from the Bayesian approach are compared to other approaches including the method of maximum likelihood or simply counting bright pixels. Uncertainty quantification (i.e., error bars) can be provided for the parameters of interest. This approach is applied to confocal images of stained biofilms of a common lab strain of Pseudomonas aeruginosa, stained biofilms of Janthinobacterium isolated from the Antarctic, and biofilms of Staphylococcusaureus that have been genetically modified to fluoresce green.

Investigating the potential use of an Antarctic variant of Janthinobacterium lividum for tackling antimicrobial resistance in a One Health approach.

The aim of this paper is to describe a new variant of Janthinobacterium lividum - ROICE173, isolated from Antarctic snow, and to investigate the antimicrobial effect of the crude bacterial extract against 200 multi-drug resistant (MDR) bacteria of both clinical and environmental origin, displaying various antibiotic resistance patterns. ROICE173 is extremotolerant, grows at high pH (5.5-9.5), in high salinity (3%) and in the presence of different xenobiotic compounds and various antibiotics. The best violacein yield (4.59 ± 0.78 mg·g-1 wet biomass) was obtained at 22 °C, on R2 broth supplemented with 1% glycerol. When the crude extract was tested for antimicrobial activity, a clear bactericidal effect was observed on 79 strains (40%), a bacteriostatic effect on 25 strains (12%) and no effect in the case of 96 strains (48%). A very good inhibitory effect was noticed against numerous MRSA, MSSA, Enterococci, and Enterobacteriaceae isolates. For several environmental E. coli strains, the bactericidal effect was encountered at a violacein concentration below of what was previously reported. A different effect (bacteriostatic vs. bactericidal) was observed in the case of Enterobacteriaceae isolated from raw vs. treated wastewater, suggesting that the wastewater treatment process may influence the susceptibility of MDR bacteria to violacein containing bacterial extracts.

Physiological and genomic insights into the lifestyle of arsenite-oxidizing Herminiimonas arsenitoxidans.

Arsenic, a representative toxic metalloid, is responsible for serious global health problems. Most organisms possess arsenic resistance strategies to mitigate this toxicity. Here, we reported a microorganism, strain AS8, from heavy metal/metalloid-contaminated soil that is able to oxidize arsenite, and investigated its physiological and genomic traits. Its cells were rod-shaped and Gram-negative, and formed small beige-pigmented colonies. 16S rRNA-based phylogenetic analysis indicated that the strain belongs to the genus Herminiimonas and is closely related to Herminiimonas glaciei UMB49T (98.7% of 16S rRNA gene sequence similarity), Herminiimonas arsenicoxydans ULPAs1T (98.4%), and Herminiimonas saxobsidens NS11T (98.4%). Under chemolithoheterotrophic conditions, the strain utilized some organic acids and amino acids as carbon and/or nitrogen sources but not electron sources. Further, the strain grew as a sulfur oxidizer in a complex medium (trypticase soy agar). Unexpectedly, most carbohydrates failed to support its growth as sole carbon sources. Genome sequencing supported these observations, and very few ABC transporters capable of oligo/monosaccharide uptake were identified in the AS8 genome. The genome harbored genes required for the colonization, flagella biosynthesis, urea degradation, and heavy metal and antibiotic resistance. Based on these polyphasic and genomic analyses, we propose that the strain AS8 be named Herminiimonas arsenitoxidans.

The effective migration of Massilia sp. WF1 by Phanerochaete chrysosporium and its phenanthrene biodegradation in soil.

Pollutant-degrading bacteria migrated by fungi may enhance the contacts between microorganisms and pollutants and improve the bioremediation efficiency of persistent organic pollutants in soil. Here, the migration of phenanthrene (PHE)-degrading bacteria Massilia sp. WF1 and Mycobacterium sp. WY10 by the hydrophobic fungi Phanerochaete chrysosporium (P. chrysosporium) and its effects on the PHE biodegradation in soil were investigated. Migration of the hydrophilic bacterium WF1 was better than that of the hydrophobic bacterium WY10 by P. chrysosporium mycelia since strain WF1 possesses flagellum and the type III secretion system. The interaction energy change of P. chrysosporium-WF1 was lower, but the interaction forces (van der Waals attractions, capillary forces, and cross-linking effects) were stronger than those of P. chrysosporium-WY10. Thus, the adhesive attraction between strain WF1 and P. chrysosporium was stronger, and consequently, strain WF1 was migrated by P. chrysosporium to a greater extent than WY10. The corresponding migration mechanism was inferred to be a bacterial 'passive' method: bacteria adhered to mycelia before they migrated with the growing mycelia. Moreover, migrated strain WF1 via P. chrysosporium showed effective PHE biodegradation in soil. Fungus-mediated migration of pollutant-degrading bacteria may play an important role in the bioremediation of pollutants in soil.

Mitigation of Membrane Biofouling in MBR Using a Cellulolytic Bacterium, Undibacterium sp. DM-1, Isolated from Activated Sludge.

Biofilm formation on the membrane surface results in the loss of permeability in membrane bioreactors (MBRs) for wastewater treatment. Studies have revealed that cellulose is not only produced by a number of bacterial species but also plays a key role during formation of their biofilm. Hence, in this study, cellulase was introduced to a MBR as a cellulose-induced biofilm control strategy. For practical application of cellulase to MBR, a cellulolytic (i.e., cellulase-producing) bacterium, Undibacterium sp. DM-1, was isolated from a lab-scale MBR for wastewater treatment. Prior to its application to MBR, it was confirmed that the cell-free supernatant of DM-1 was capable of inhibiting biofilm formation and of detaching the mature biofilm of activated sludge and cellulose-producing bacteria. This suggested that cellulase could be an effective anti-biofouling agent for MBRs used in wastewater treatment. Undibacterium sp. DM-1-entrapping beads (i.e., cellulolytic-beads) were applied to a continuous MBR to mitigate membrane biofouling 2.2-fold, compared with an MBR with vacant-beads as a control. Subsequent analysis of the cellulose content in the biofilm formed on the membrane surface revealed that this mitigation was associated with an approximately 30% reduction in cellulose by cellulolytic-beads in MBR.

Oxalic acid: a signal molecule for fungus-feeding bacteria of the genus Collimonas?

Mycophagous (=fungus feeding) soil bacteria of the genus Collimonas have been shown to colonize and grow on hyphae of different fungal hosts as the only source of energy and carbon. The ability to exploit fungal nutrient resources might require a strategy for collimonads to sense fungi in the soil matrix. Oxalic acid is ubiquitously secreted by soil fungi, serving different purposes. In this study, we investigated the possibility that collimonads might use oxalic acid secretion to localize a fungal host and move towards it. We first confirmed earlier indications that collimonads have a very limited ability to use oxalic acid as growth substrate. In a second step, with using different assays, we show that oxalic acid triggers bacterial movement in such a way that accumulation of cells can be expected at micro-sites with high free oxalic acid concentrations. Based on these observations we propose that oxalic acid functions as a signal molecule to guide collimonads to hyphal tips, the mycelial zones that are most sensitive for mycophagous bacterial attack.

The cutaneous bacterium Janthinobacterium lividum inhibits the growth of Trichophyton rubrum in vitro.

BACKGROUND: Tinea pedis (athlete's foot) is a fungal infection that is both widespread and challenging to treat. Standard treatments consist of topical and systemic therapies of antifungal agents, such as miconazole, itraconazole, and terbinafine. The extended nature of topical therapy and the toxicity of long-term systemic therapy limit the utility of current treatments. An alternate approach relies on an understanding of bacterial-fungal interactions. Specifically, a probiotic antifungal bacterium such as Janthinobacterium lividum can counter infection; Janthinobacterium is a major constituent of the human skin microbiota. Janthinobacterium lividum has been shown to ameliorate the effects of the cutaneous fungal disease chytridiomycosis in a vertebrate species (Rana muscosa). METHODS: Dual-culture plate challenge assays were performed using J. lividum and Trichophyton rubrum, the leading cause of athlete's foot. RESULTS: In all cases, T. rubrum colonies grew significantly smaller when co-cultured with J. lividum. CONCLUSION: These in vitro results suggest that J. lividum merits further investigation as a human cutaneous probiotic.

Massilia kyonggiensis sp. nov., isolated from forest soil in Korea.

A Gram-negative, short, rod-shaped bacterium, TSA1(T), was isolated from forest soil collected at Kyonggi University, South Korea. Assessment of 16S rRNA gene sequence similarity indicated that the strain is related to Massilia niastensis 5516S-1(T) (98.3%), M. haematophila CCUG 38318(T) (97.9%), M. aerilata 5516S-11(T) (97.9%), M. tieshanensis TS3(T) (97.6%), and M. varians CCUG 3529(T) (97.1%). Colonies grown on Reasoner's 2A agar at 30°C for 2 days were transparent, white, round, smooth, and glossy. The cells grew at 10-42°C (optimum: 25-37°C) and pH 5-9 (optimum: 5-9) and in 0-2% NaCl (optimum: 0-1%). TSA1(T) was able to grow on trypticase soy and nutrient agar, but not on Luria-Bertani or MacConkey agar. The strain was catalase- and oxidasepositive and able to degrade starch and casein, but not carboxymethyl cellulose. The predominant quinone of TSA1(T) was Q-8, the major fatty acids were summed feature 3 and C16:0, and the DNA G+C content was 66.7 mol%. Given these findings, we propose that this strain is a novel species of the genus Massilia. We suggest the name Massilia kyonggiensis sp. nov. (type strain, KACC 17471(T) =KEMB 9005-031(T) =JCM 19189(T)).

Influence of hydraulic regimes on bacterial community structure and composition in an experimental drinking water distribution system.

Microbial biofilms formed on the inner-pipe surfaces of drinking water distribution systems (DWDS) can alter drinking water quality, particularly if they are mechanically detached from the pipe wall to the bulk water, such as due to changes in hydraulic conditions. Results are presented here from applying 454 pyrosequencing of the 16S ribosomal RNA (rRNA) gene to investigate the influence of different hydrological regimes on bacterial community structure and to study the potential mobilisation of material from the pipe walls to the network using a full scale, temperature-controlled experimental pipeline facility accurately representative of live DWDS. Analysis of pyrosequencing and water physico-chemical data showed that habitat type (water vs. biofilm) and hydraulic conditions influenced bacterial community structure and composition in our experimental DWDS. Bacterial community composition clearly differed between biofilms and bulk water samples. Gammaproteobacteria and Betaproteobacteria were the most abundant phyla in biofilms while Alphaproteobacteria was predominant in bulk water samples. This suggests that bacteria inhabiting biofilms, predominantly species belonging to genera Pseudomonas, Zooglea and Janthinobacterium, have an enhanced ability to express extracellular polymeric substances to adhere to surfaces and to favour co-aggregation between cells than those found in the bulk water. Highest species richness and diversity were detected in 28 days old biofilms with this being accentuated at highly varied flow conditions. Flushing altered the pipe-wall bacterial community structure but did not completely remove bacteria from the pipe walls, particularly under highly varied flow conditions, suggesting that under these conditions more compact biofilms were generated. This research brings new knowledge regarding the influence of different hydraulic regimes on the composition and structure of bacterial communities within DWDS and the implication that this might have on drinking water quality.

Massilia flava sp. nov., isolated from soil.

A Gram-stain-negative, rod-shaped bacterium, designated strain Y9(T), was isolated from a soil sample collected in Ningxia Province in China and was characterized to determine its taxonomic position. Strain Y9(T) contained Q-8 as the predominant ubiquinone. Major fatty acid components were summed feature 3 (C(16:1)ω7c and/or iso-C(15:0) 2-OH) and C(16:0). The major polar lipids were phosphatidylethanolamine, phosphatidylglycerol and diphosphatidylglycerol. The G+C content of the genomic DNA of strain Y9(T) was 68.7 mol%. A phylogenetic tree based on 16S rRNA gene sequences showed that the strain fell within the evolutionary radiation encompassed by the genus Massilia. Levels of 16S rRNA gene sequence similarity between strain Y9(T) and the type strains of recognized Massilia species ranged from 95.2 to 98.2%, the highest values being with Massilia albidiflava 45(T) (98.2%) and Massilia lutea 101(T) (98.0%). However, levels of DNA-DNA relatedness between strain Y9(T) and M. albidiflava KCTC 12343(T) and M. lutea KCTC 12345(T) were 37 and 26%, respectively. Strain Y9(T) was clearly differentiated from its nearest phylogenetic relatives in the genus Massilia based on phenotypic, chemotaxonomic and phylogenetic properties. Therefore, strain Y9(T) is considered to represent a novel species of the genus Massilia, for which the name Massilia flava sp. nov. is proposed. The type strain is Y9(T) (=CGMCC 1.10685(T) =KCTC 23585(T)).

Geosmin degradation by seasonal biofilm from a biological treatment facility.

INTRODUCTION: Initial geosmin degradation was closely related to water temperature and natural geosmin concentration of sampling environment. Here, for the first time, we evaluated the biodegradation of geosmin by microorganisms in biofilm from biological treatment unit of actual potable water treatment plant. MATERIALS AND METHODS: At an initial geosmin concentration of 2,500 ng/l, efficient geosmin removal was confirmed throughout the year. Furthermore, in the presence of mixed musty odor compounds (geosmin and MIB) as carbon source, geosmin degradation was enhanced compared to sole carbon source (geosmin alone). RESULTS AND DISCUSSION: PCR-DGGE analysis revealed a rich community structure within the biofilm during rapid geosmin removal period, April. PCA revealed that the significant change in bacterial communities occurred from day 1 to day 2. Two novel geosmin-degrading bacteria were isolated from the biofilm of the biological treatment unit of Kasumigaura Water Purification, Waterworks Department, Japan. They belong to Methylobacterium sp. and Oxalobacteraceae bacterium, respectively. CONCLUSIONS: These studies provide further insights into the unknown microbiological processes that occur during the biological removal of geosmin through water treatment and could facilitate the geosmin bioremediation in contaminated habitats.