0000000001238267
AUTHOR
Marja Tiirola
Influence of rearing conditions on Flavobacterium columnare infection of rainbow trout, Oncorhynchus mykiss (Walbaum).
The influence of rearing conditions on Flavobacterium columnare infection of rainbow trout, Oncorhynchus mykiss (Walbaum), was studied experimentally in the laboratory and at a fish farm. In experiment I, the effect of parasitic infection on columnaris disease was studied using F. columnare carrier fish. The fish were exposed to Diplostomum spathaceum cercariae and a set of other stressors in order to induce clinical columnaris infection. Parasitic infection and other stressors failed to induce the disease. Disease occurred when the fish were challenged with F. columnare, but D. spathaceum infection did not enhance the severity of the infection. In experiment II, the influence of rearing de…
Suppression of Saprolegnia infections in rainbow trout (Oncorhynchus mykiss) eggs using protective bacteria and ultraviolet irradiation of the hatchery water
Since formalin is widely used in prevention of Sapro- legnia infections in salmonid fish hatcheries, there is a need for more environmentally safe treatment methods. Therefore, we screened 360 bacterial iso- lates against their ability to antagonize the growth of Saprolegnia parasitica hyphae in vitro, and best strains were selected according to their antagonistic proper- ties and colonization capability on rainbow trout egg surface. Protective bacterial cultures of Pseudomonas sp. M162, Pseudomonas sp. M174 and Janthinobacte- rium M169 were tested for prevention of Saprolegnia sp. infections during incubation trials of rainbow trout (Oncorhynchus mykiss )e ggs with UV irradiated (400 mWs c…
Whole-lake dissolved inorganic 13C additions reveal seasonal shifts in zooplankton diet.
Sustained whole-lake additions of 13C-enriched dissolved inorganic carbon (DIC), intended to increase experimentally the delta13C of DIC in the epilimnion of a small lake with high dissolved organic carbon (DOC), were made during three seasonal periods (spring, summer, and autumn). Coupled with carbon and nitrogen stable isotope analysis of zooplankton and several of their putative food sources, these additions were used to investigate seasonal changes in the relative contributions of different food sources to zooplankton diet in the lake. Four main potential food sources were considered: phytoplankton, heterotrophic bacteria (HB), methanotrophic bacteria (MOB), and green sulfur bacteria (G…
Prebiotic Xylo-Oligosaccharides Ameliorate High-Fat-Diet-Induced Hepatic Steatosis in Rats
Understanding the importance of the gut microbiota (GM) in non-alcoholic fatty liver disease (NAFLD) has raised the hope for therapeutic microbes. We have shown that high hepatic fat content associated with low abundance of Faecalibacterium prausnitzii in humans and, further, the administration of F. prausnitzii prevented NAFLD in mice. Here, we aimed at targeting F. prausnitzii by prebiotic xylo-oligosaccharides (XOS) to treat NAFLD. First, the effect of XOS on F. prausnitzii growth was assessed in vitro. Then, XOS was supplemented or not with high (HFD, 60% of energy from fat) or low (LFD) fat diet for 12 weeks in Wistar rats (n = 10/group). XOS increased F. prausnitzii growth, having onl…
Production and characterization of the recombinant Sphingomonas chlorophenolica pentachlorophenol 4-monooxygenase.
Abstract Pentachlorophenol 4-monooxygenase (PCP4MO) from Sphingomonas chlorophenolica is a flavoprotein that hydroxylates PCP in the presence of NADPH and oxygen. In order to investigate the structure and function of active site, recombinant PCP4MO (rePCP4MO) was produced in Escherichia coli as a glutathione S-transferase (GST) fusion protein. Moreover, a tobacco etch virus (TEV) protease cleavage site (EKLYFQG) was introduced into GST-PCP4MO and a his-tagged TEV protease was employed. Hence, a two-step purification protocol was developed which allowed obtaining 15–20 mg of rePCP4MO from 1 L culture. The rePCP4MO revealed identity with native enzyme by SDS–PAGE and N-terminal sequence analy…
Genetic and Environmental Controls on Nitrous Oxide Accumulation in Lakes
We studied potential links between environmental factors, nitrous oxide (N2O) accumulation, and genetic indicators of nitrite and N2O reducing bacteria in 12 boreal lakes. Denitrifying bacteria were investigated by quantifying genes encoding nitrite and N2O reductases (nirS/nirK and nosZ, respectively, including the two phylogenetically distinct clades nosZ(I) and nosZ(II)) in lake sediments. Summertime N2O accumulation and hypolimnetic nitrate concentrations were positively correlated both at the inter-lake scale and within a depth transect of an individual lake (Lake Vanajavesi). The variability in the individual nirS, nirK, nosZ(I), and nosZ(II) gene abundances was high (up to tenfold) a…
Freezing Induces Biased Results in the Molecular Detection of Flavobacterium columnare
ABSTRACT Specific PCR detection and electron microscopy of Flavobacterium columnare revealed the risk of false-negative results in molecular detection of this fish pathogen. Freezing and thawing destroyed the cells so that DNA was for the most part undetectable by PCR. The detection of bacteria was also weakened after prolonged enrichment cultivation of samples from infected fish.
The role ofSphagnummosses in the methane cycling of a boreal mire
Peatlands are a major natural source of atmospheric methane (CH4). Emissions from Sphagnum-dominated mires are lower than those measured from other mire types. This observation may partly be due to methanotrophic (i.e., methane-consuming) bacteria associated with Sphagnum. Twenty-three of the 41 Sphagnum species in Finland can be found in the peatland at Lakkasuo. To better understand the Sphagnum-methanotroph system, we tested the following hypotheses: (1) all these Sphagnum species support methanotrophic bacteria; (2) water level is the key environmental determinant for differences in methanotrophy across habitats; (3) under dry conditions, Sphagnum species will not host methanotrophic ba…
Microbiome response to foam fractionation and ozonation in RAS
Efficient water treatment is required to maintain high water quality and control microbial growth in recirculating aquaculture systems (RAS). Here, we examined the effects of two treatment methods, ozonation and foam fractionation, separately and combined, on the microbiology in twelve identical experimental RAS with rainbow trout (Oncorhyncus mykiss) during 8 weeks. Microbes suspended in water and growing in biofilter biofilms were examined using flow cytometry analysis and high throughput sequencing of the 16S rRNA gene. The results showed that foam fractionation did not cause large changes in abundance or overall community composition of free-living microbes. Instead, through decreasing …
Bacterial communities in Arctic fjelds of Finnish Lapland are stable but highly pH-dependent
The seasonal and spatial variations of microbial communities in Arctic fjelds of Finnish Lapland were studied. Phospholipid fatty acid analysis (PLFA) and terminal restriction fragment analysis (T-RFLP) of amplified 16S rRNA genes were used to assess the effect of soil conditions and vegetation on microbial community structures along different altitudes of two fjelds, Saana and Jehkas. Terminal restriction fragments were additionally analysed from c . 160 cloned sequences and isolated bacterial strains and matched with those of soil DNA samples. T-RFLP and PLFA analyses indicated relatively similar microbial communities at various altitudes and under different vegetation of the two fjelds. …
Methylophaga and Hyphomicrobium can be used as target genera in monitoring saline water methanol-utilizing denitrification.
Abstract Which bacterial taxonomic groups can be used in monitoring saline water methanol-utilizing denitrification and whether nitrate is transformed into N2 in the process are unclear. Therefore, methylotrophic bacterial communities of two efficiently functioning (nitrate/nitrite reduction was 63–96 %) tropical and cool seawater reactors at a public aquarium were investigated with clone library analysis and 454 pyrosequencing of the 16S rRNA genes. Transformation of nitrate into N2 was confirmed using 15N labeling in incubation of carrier material from the tropical reactor. Combining the data with previous study results, Methylophaga and Hyphomicrobium were determined to be suitable targe…
Methane-oxidizing and photoautotrophic bacteria are major producers in a humic lake with large anoxic hypolimnion
Heterotrophic processes are considered to prevail in lakes with high terrestrial loading and a high content of dissolved organic carbon. However, previous studies have shown abundant methanotrophic bacteria (MOB) and photoautotrophic green sulphur bacteria (GSB) in some boreal lakes. We used genetic and lipid biomarkers to follow the seasonal succession of the abundance and carbon biomass of the major bacterial groups in Lake Mekkojarvi, a small Finnish polyhumic lake with a large anoxic hypolimnion. Biomarkers were based on length heterogeneity analysis of 16S rRNA gene amplification by polymerase chain reaction (PCR) (LH-PCR) and on phospholipid fatty acid (PLFA) analysis. Representative …
Effect of peracetic acid on levels of geosmin, 2-methylisoborneol, and their potential producers in a recirculating aquaculture system for rearing rainbow trout (Oncorhynchus mykiss)
In recirculating aquaculture systems (RAS)s, off-flavors and odors, mainly caused by geosmin (GSM) and 2-methylisoborneol (MIB), can accumulate in the flesh of fish from RAS water, reducing the profitability of production. In this study, peracetic acid (PAA) was applied in three application intervals to pump sumps of rainbow trout (Oncorhynchus mykiss) reared in RAS. Using a real-time polymerase chain reaction (qPCR), the potential off-flavor producers were quantified using geoA and MIB synthase genes. Streptomyces was identified as the major GSM producer, and biofilters showed the highest number of potential off-flavor producers. Concentrations of GSM and MIB were analyzed in the circulati…
Interlake variation and environmental controls of denitrification across different geographical scales
Denitrification in lakes significantly reduces the nitrogen (N) load from land to oceans, but the factors controlling it remain poorly under- stood. Therefore, interlake variation of denitrification in sediments of small to medium-sized lakes (from 0.03 to 17.8 km 2 ) was studied across different geo- graphical scales. At the local scale, the denitrification rates and sediment microbial communities were studied in 4 boreal lakes in close proximity (within 20 km) using the isotope pairing technique (IPT) and molecular methods. These local scale data were com- bined with previously published data on denitrifica- tion rates from 10 other European lakes to extend the analysis to the regional (b…
Mucilaginibacter frigoritolerans sp. nov., Mucilaginibacter lappiensis sp. nov. and Mucilaginibacter mallensis sp. nov., isolated from soil and lichen samples.
Five cold-adapted bacteria belonging to the genus Mucilaginibacter were isolated from lichen and soil samples collected from Finnish Lapland and investigated in detail by phenotypic and phylogenetic analyses. Based on 16S rRNA gene phylogeny, the novel strains represent three new branches within the genus Mucilaginibacter. The strains were aerobic, chemo-organotrophic, non-motile rods and formed pigmented, smooth, mucoid colonies on solid media. The strains grew between 0 and 33 °C (optimum growth at 25 °C) and at pH 4.5–8.0 (optimum growth at pH 6.0). The main cellular fatty acids were iso-C15 : 0, summed feature 3 (C16 : 1 ω7c/iso-C15 : 0 2-OH) and iso-C17 : 0 3-OH and the major respirato…
The Value of Liquid Biopsies for Guiding Therapy Decisions in Non-small Cell Lung Cancer
Targeted therapies have allowed for an individualized treatment approach in non-small-cell lung cancer (NSCLC). The initial therapeutic decisions and success of targeted therapy depend on genetic identification of personal tumor profiles. Tissue biopsy is the gold standard for molecular analysis, but non-invasive or minimally invasive liquid biopsy methods are also now used in clinical practice, allowing for later monitoring and optimization of the cancer treatment. The inclusion of liquid biopsy in the management of NSCLC provides strong evidence on early treatment response, which becomes a basis for determining disease progression and the need for changes in treatment. Liquid biopsies can…
The effect of iron on the biodegradation of natural dissolved organic matter
Iron (Fe) may alter the biodegradation of dissolved organic matter (DOM), by interacting with (DOM), phosphorus (P) and microbes. We isolated DOM and a bacterial community from boreal lake water and examined bacterial growth on DOM in laboratory experiments. Fe was introduced either together with DOM (DOM-Fe) or into bacterial suspension, which led to the formation of insoluble Fe-precipitates on bacterial surfaces (Fe coating). In the latter case, the density of planktonic bacteria was an order of magnitude lower than that in the corresponding treatment without introduced Fe. The association of Fe with DOM decreased bacterial growth, respiration, and growth efficiency compared with DOM alo…
Gammaproteobacterial methanotrophs dominate methanotrophy in aerobic and anaerobic layers of boreal lake waters
Small oxygen-stratified humic lakes of the boreal zone are important sources of methane to the atmosphere. Although stable isotope profiling has indicated that a substantial part of methane is already oxidized in the anaerobic water layers in these lakes, the contributions of aerobic and anaerobic methanotrophs in the process are unknown. We used next-generation sequencing of mcrA and 16S rRNA genes to characterize the microbial communities in the water columns of 2 boreal lakes in Finland, Lake Alinen-Mustajärvi and Lake Mekkojärvi, and complemented this with a shotgun metagenomic analysis from Alinen-Mustajärvi and an analysis of pmoA genes and 16S rRNA, mcrA, and pmoA transcripts from Me…
Salinity affects nitrate removal and microbial composition of denitrifying woodchip bioreactors treating recirculating aquaculture system effluents
This study investigated the effect of salinity on microbial composition and denitrification capacity of woodchip bioreactors treating recirculating aquaculture system (RAS) effluents. Twelve laboratory-scale woodchip bioreactors were run in triplicates at 0, 15, 25, and 35 ppt salinities, and water chemistry was monitored every third day during the first 39 days of operation. Microbial communities of the woodchips bioreactors were analyzed at the start, after one week, and at the end of the trial. Woodchip bioreactors removed nitrate at all salinities tested. The highest NO3-N removal rate of 22.0 ± 6.9 g NO3-N/m3/d was obtained at 0 ppt, while 15.3 ± 4.9, 12.5 ± 5.4 and 11.8 ± 4.0 g NO3-N/…
Nitrogen fixation and methanotrophy in forest mosses along a N deposition gradient
Abstract Nitrogen deposition has decreased the plant-associated nitrogen (N 2 ) fixation when measured using the indirect acetylene reduction assay (ARA). However, nitrogen deposition can also lead to changes in the diversity of moss symbionts, e.g. affect methanotrophic N 2 fixation, which is not measured by ARA. To test this hypothesis we compared ARA with the direct stable isotope method ( 15 N 2 incorporation) and studied methanotrophy in two mosses, Hylocomium splendens and Pleurozium schreberi , collected from seven forest sites along a boreal latitudinal N deposition transect. We recognized that the two independent N 2 fixation measures gave corresponding results with the conversion …
Novosphingobium lentum sp. nov., a psychrotolerant bacterium from a polychlorophenol bioremediation process
A polychlorophenol-degrading strain, designated MT1T, and three MT1-like strains, MT101, MT103 and MT104, were isolated from a cold (4–8 °C) fluidized-bed process treating chlorophenol-contaminated groundwater in southern Finland. The organisms were Gram-negative, rod-shaped, catalase-positive, non-spore-forming and non-motile. Phylogenetic analysis based on 16S rRNA gene sequences indicated that the strains belonged to the α-4 subclass of the Proteobacteria and were members of the genus Novosphingobium. The highest 16S rRNA gene sequence similarity observed for these strains was 96·5 % with the type strains of Novosphingobium hassiacum, Novosphingobium aromaticivorans and Novosphingobium s…
Sphingomonas fennica sp. nov. and Sphingomonas haloaromaticamans sp. nov., outliers of the genus Sphingomonas.
Bacterial isolates obtained from polychlorophenol-contaminated sites in Finland (strain K101T) and from a Dutch drinking water well (strain A175T) were characterized taxonomically. 16S rRNA gene sequence analysis, determination of DNA G+C content, physiological characterization, estimation of the ubiquinone and polar lipid patterns and fatty acid content revealed that strains K101T and A175T were similar to Sphingomonas wittichii RW1T but also showed pronounced differences. The DNA G+C contents of the two novel strains were 63.6 and 66.1 mol%, respectively. On the basis of these results, two novel species of the genus Sphingomonas are described, for which the names Sphingomonas haloaromatic…
Effects of alternative electron acceptors on the activity and community structure of methane-producing and consuming microbes in the sediments of two shallow boreal lakes
The role of anaerobic CH4 oxidation in controlling lake sediment CH4 emissions remains unclear. Therefore, we tested how relevant EAs (SO42−, NO3−, Fe3+, Mn4+, O2) affect CH4 production and oxidation in the sediments of two shallow boreal lakes. The changes induced to microbial communities by the addition of Fe3+ and Mn4+ were studied using next-generation sequencing targeting the 16S rRNA and methyl-coenzyme M reductase (mcrA) genes and mcrA transcripts. Putative anaerobic CH4-oxidizing archaea (ANME-2D) and bacteria (NC 10) were scarce (up to 3.4% and 0.5% of archaeal and bacterial 16S rRNA genes, respectively), likely due to the low environmental stability associated with shallow depths.…
Biofiltration of airborne VOCs with green wall systems : microbial and chemical dynamics
Botanical air filtration is a promising technology for reducing indoor air contaminants, but the underlying mechanisms need better understanding. Here, we made a set of chamber fumigation experiments of up to 16 weeks of duration, to study the filtration efficiencies for seven volatile organic compounds (VOCs; decane, toluene, 2-ethylhexanol, α-pinene, octane, benzene, and xylene) and to monitor microbial dynamics in simulated green wall systems. Biofiltration functioned on sub-ppm VOC levels without concentration-dependence. Airflow through the growth medium was needed for efficient removal of chemically diverse VOCs, and the use of optimized commercial growth medium further improved the e…
Microbial communities in full-scale woodchip bioreactors treating aquaculture effluents.
Woodchip bioreactors are being successfully applied to remove nitrate from commercial land-based recirculating aquaculture system (RAS) effluents. In order to understand and optimize the overall function of these bioreactors, knowledge on the microbial communities, especially on the microbes with potential for production or mitigation of harmful substances (e.g. hydrogen sulfide; H2S) is needed. In this study, we quantified and characterized bacterial and fungal communities, including potential H2S producers and consumers, using qPCR and high throughput sequencing of 16S rRNA gene. We took water samples from bioreactors and their inlet and outlet, and sampled biofilms growing on woodchips a…
The effects of different combinations of fixed and moving bed bioreactors on rainbow trout (Oncorhynchus mykiss) growth and health, water quality and nitrification in recirculating aquaculture systems
The effect of bioreactor design on nitrification efficiency has been well studied, but less is known about the overall impacts on water quality. Besides nitrification, submerged fixed bed bioreactors (FBBR) trap fine solid particles, whereas moving bed bioreactors (MBBR) grind solids, possibly increasing solids and particle accumulation in the system. In this experiment, the effects of different combinations of fixed bed and moving bed bioreactors on water quality, solids removal, particle size distribution, fish health based on histopathological changes and nitrification efficiency were studied in laboratory scale recirculating aquaculture systems (RAS) with rainbow trout (Oncorhynchus myk…
Vertical stratification of bacteria and archaea in sediments of a small boreal humic lake
ABSTRACT Although sediments of small boreal humic lakes are important carbon stores and greenhouse gas sources, the composition and structuring mechanisms of their microbial communities have remained understudied. We analyzed the vertical profiles of microbial biomass indicators (PLFAs, DNA and RNA) and the bacterial and archaeal community composition (sequencing of 16S rRNA gene amplicons and qPCR of mcrA) in sediment cores collected from a typical small boreal lake. While microbial biomass decreased with sediment depth, viable microbes (RNA and PLFA) were present all through the profiles. The vertical stratification patterns of the bacterial and archaeal communities resembled those in mar…
Acidobacteria dominate the active bacterial communities of Arctic tundra with widely divergent winter-time snow accumulation and soil temperatures.
The timing and extent of snow cover is a major controller of soil temperature and hence winter-time microbial activity and plant diversity in Arctic tundra ecosystems. To understand how snow dynamics shape the bacterial communities, we analyzed the bacterial community composition of windswept and snow-accumulating shrub-dominated tundra heaths of northern Finland using DNA- and RNA-based 16S rRNA gene community fingerprinting (terminal restriction fragment polymorphism) and clone library analysis. Members of the Acidobacteria and Proteobacteria dominated the bacterial communities of both windswept and snow-accumulating habitats with the most abundant phylotypes corresponding to subdivision …
Resistant ammonia-oxidizing archaea endure, but adapting ammonia-oxidizing bacteria thrive in boreal lake sediments receiving nutrient-rich effluents
Climate change along with anthropogenic activities changes biogeochemical conditions in lake ecosystems, modifying the sediment microbial communities. Wastewater effluents introduce nutrients and organic material but also novel microbes to lake ecosystems, simulating forthcoming increases in catchment loadings. In this work, we first used 16s rRNA gene sequencing to study how the overall sediment microbial community responds to wastewater in six boreal lakes. To examine forthcoming changes in the lake biogeochemistry, we focused on the ammonia-oxidizing archaea (AOA) and bacteria (AOB), and examined their functional and compositional community response to wastewater. Although we found the l…
Diagnosis of flavobacteriosis by direct amplification of rRNA genes
A broad-range bacterial PCR method with universal 16S rDNA targeting primers and bacterial cultivation was used to identify the putative pathogen in flavobacterial outbreaks. Restriction fragment length polymorphism (PCR-RFLP) analysis and sequencing of the partial 16S rDNA PCR products of 10 skin samples and 10 representative isolates derived from the same fish specimens revealed differences between direct molecular and cultivation-based analysis. Flavobacterium columnare-like sequences dominated in the direct molecular analysis in most cases, whereas most of the isolates belonged to a phylogenetically heterogeneous group of flavobacteria clustering with F. hibernum. F. columnare was isola…
Sample Preservation, DNA or RNA Extraction and Data Analysis for High-Throughput Phytoplankton Community Sequencing
Phytoplankton is the basis for aquatic food webs and mirrors the water quality. Conventionally, phytoplankton analysis has been done using time consuming and partly subjective microscopic observations, but next generation sequencing (NGS) technologies provide promising potential for rapid automated examination of environmental samples. Because many phytoplankton species have tough cell walls, methods for cell lysis and DNA or RNA isolation need to be efficient to allow unbiased nucleic acid retrieval. Here, we analyzed how two phytoplankton preservation methods, three commercial DNA extraction kits and their improvements, three RNA extraction methods, and two data analysis procedures affect…
Cooperation between CRISPR-Cas types enables adaptation in an RNA-targeting system
AbstractCRISPR-Cas immune systems adapt to new threats by acquiring spacers from invading nucleic acids such as phage genomes. However, some CRISPR-Cas loci lack genes necessary for spacer acquisition, despite apparent variation in spacer content between strains. It has been suggested that such loci may use acquisition machinery from co-occurring CRISPR-Cas systems. Here, using a lytic dsDNA phage, we observe spacer acquisition in the native host Flavobacterium columnare that carries an acquisition-deficient subtype VI-B locus and a complete subtype II-C locus. We characterize acquisition events in both loci and show that the RNA-targeting VI-B locus acquires spacers in trans using acquisit…
Methylophilaceae and Hyphomicrobium as target taxonomic groups in monitoring the function of methanol-fed denitrification biofilters in municipal wastewater treatment plants
Abstract Molecular monitoring of bacterial communities can explain and predict the stability of bioprocesses in varying physicochemical conditions. To study methanol-fed denitrification biofilters of municipal wastewater treatment plants, bacterial communities of two full-scale biofilters were compared through fingerprinting and sequencing of the 16S rRNA genes. Additionally, 16S rRNA gene fingerprinting was used for 10-week temporal monitoring of the bacterial community in one of the biofilters. Combining the data with previous study results, the family Methylophilaceae and genus Hyphomicrobium were determined as suitable target groups for monitoring. An increase in the relative abundance …
Chondroitin AC lyase activity is related to virulence of fish pathogenic Flavobacterium columnare.
The virulence of eight Flavobacterium columnare strains was studied to find correlations between several virulence-related factors and virulence. Virulence was tested in vivo using rainbow trout, Oncorhynchus mykiss (Walbaum). Suggested virulence-related factors such as production of the degradative enzyme chondroitin lyase, plasmid occurrence and adhesion capability were studied in vitro. Infection with the four most virulent strains resulted in 95-100% mortality within 114 h. Chondroitin lyase activity was found to be significantly related to the virulence of the strains at 25 degrees C and it was also shown to be temperature-dependent, being higher at 25 degrees C than at 20 degrees C. V…
Spatial and temporal variation in denitrification and in the denitrifier community in a boreal lake
We investigated the spatial and temporal variation in denitrification rates (isotopepairing technique, IPT) and in the denitrifier community (examination of gene nirK by denaturinggradient gel electrophoresis [DGGE] of microbial DNA) in the sediments of a boreal, clear-water, eutrophic lake using samples collected from shallow littoral, deep littoral and shallow profundal sediments during early summer, mid-summer, autumn and winter. The measured denitrification rates (44 to 613 μmol N m−2 d−1) are among the lowest ever reported from lacustrine sediments. Denitri fi cation rates varied both spatially and temporally, being highest in the profundal zone during midsummer and in the littoral zon…
Reliability of mitochondrial DNA in an acanthocephalan: The problem of pseudogenes
The utility of mitochondrial DNA as a molecular marker for evolutionary studies is well recognized. However, several problems can arise when using mitochondrial DNA, one of which is the presence of nuclear mitochondrial pseudogenes, or Numts. Pseudogenes of cytochrome oxidase I were preferentially amplified from Acanthocephalus lucii (Acanthocephala) using a universal PCR approach. To verify the presence and abundance of pseudogenes, length heterogeneity analysis of the PCR fragments was performed. PCR products obtained with universal primers often contained fragments of different sizes. Cloned sequences from universal PCR products nearly always contained sequence abnormalities such as inde…
Cryptogams signify key transition of bacteria and fungi in Arctic sand dune succession
SummaryPrimary succession models focus on aboveground vascular plants. However, the prevalence of mosses and lichens, i.e. cryptogams, suggests they play a role in soil successions. Here, we explore whether effects of cryptogams on belowground microbes can facilitate progressive shifts in sand dune succession.We linked aboveground vegetation, belowground bacterial and fungal community, and soil chemistry in six successional stages in Arctic inland sand dunes: bare sand, grass, moss, lichen, ericoid heath and mountain birch forest.Compared to the bare sand and grass stages, microbial biomass and the proportion of fungi increased in the moss stage, and later stage microbial groups appeared de…
Directional high-throughput sequencing of RNAs without gene-specific primers.
Ribosomal RNA analysis is a useful tool for characterization of microbial communities. However, the lack of broad-range primers has hampered the simultaneous analysis of eukaryotic and prokaryotic members by amplicon sequencing. We present a complete workflow for directional, primer-independent sequencing of size-selected small subunit ribosomal RNA fragments. The library preparation protocol includes gel extraction of the target RNA, ligation of an RNA oligo to the 5′-end of the target, and cDNA synthesis with a tailed random-hexamer primer and further barcoding. The sequencing results of a phytoplankton mock community showed a highly similar profile to the biomass indicators. This method…
The effect of peracetic acid on microbial community, water quality, nitrification and rainbow trout (Oncorhynchus mykiss) performance in recirculating aquaculture systems
Abstract Microbial biofilters control water quality and enable the overall function of recirculation aquaculture systems (RAS). Changes in environmental conditions can affect the abundance and interactions of the diverse microbial populations of the biofilter, affecting nitrification of harmful ammonium and thus fish health. Here, we examined the effect of different application frequencies (0, 1, 2 and 4 times per week) of a common disinfectant, peracetic acid (PAA, applied 1.1 mg l−1 twice per day), on biofilter microbial communities, focusing especially on nitrifying microbial groups and using a high throughput sequencing of 16S rRNA gene and quantitative PCR (qPCR). In addition, we measu…
Functional gene pyrosequencing reveals core proteobacterial denitrifiers in boreal lakes
Denitrification is an important microbial process in aquatic ecosystems that can reduce the effects of eutrophication. Here, quantification and pyrosequencing of nirS, nirK, and nosZ genes encoding for nitrite and nitrous oxide reductases was performed in sediment samples from four boreal lakes to determine the structure and seasonal stability of denitrifying microbial populations. Sediment quality and nitrate concentrations were linked to the quantity and diversity of denitrification genes, the abundance of denitrifying populations (nirS and nosZ genes) correlated with coupled nitrificationdenitrification (Dn), and the denitrification of the overlying water NO3 − (Dw) correlated with the n…
The effects of different combinations of fixed and moving bed bioreactors on rainbow trout (Oncorhynchus mykiss) growth and health, water quality and nitrification in recirculating aquaculture systems
Abstract The effect of bioreactor design on nitrification efficiency has been well studied, but less is known about the overall impacts on water quality. Besides nitrification, submerged fixed bed bioreactors (FBBR) trap fine solid particles, whereas moving bed bioreactors (MBBR) grind solids, possibly increasing solids and particle accumulation in the system. In this experiment, the effects of different combinations of fixed bed and moving bed bioreactors on water quality, solids removal, particle size distribution, fish health based on histopathological changes and nitrification efficiency were studied in laboratory scale recirculating aquaculture systems (RAS) with rainbow trout (Oncorhy…
Out of sight : Profiling soil characteristics, nutrients and bacterial communities affected by organic amendments down to one meter in a long-term maize experiment
Common soil characteristics, nutrients and microbial activity at deeper soil depths are topics seldom covered in agricultural studies. Biogeochemical cycles in deep soils are not yet fully understood. This study investigates the effect of different mineral and organic fertilisation on soil organic matter dynamics, nutrients and bacterial community composition in the first meter of the soil profiles in the long-term maize cropping system experiment Tetto Frati, near the Po River in northern Italy. The following treatments have been applied since 1992: 1) crop residue removal (CRR), 2) crop residue incorporation (CRI), 3) crop residue removal with bovine slurry fertilisation (SLU), 4) crop re…
Construction of Metatranscriptomic Libraries for 5′ End Sequencing of rRNAs for Microbiome Research
Metatranscriptomic sequencing enables studying community-wide gene expression profiles of microbial samples and getting functional insight on their up- or downregulated pathways. However, shotgun sequencing is not the most efficient way to study expression of ribosomal RNA genes or to compare lot of samples in experimental setups. Here we describe an efficient primer-independent method for processing and barcoding libraries for directional sequencing of the 5' end region of the RNA. When applying size selection of the original RNA, the method forms an optimal solution for the simultaneous analysis of bacterial, archaeal, and eukaryotic rRNA diversity.
Enhanced nitrogen removal of low carbon wastewater in denitrification bioreactors by utilizing industrial waste toward circular economy
Aquaculture needs practical solutions for nutrient removal to achieve sustainable fish production. Passive denitrifying bioreactors may provide an ecological, low-cost and low-maintenance approach for wastewater nitrogen removal. However, innovative organic materials are needed to enhance nitrate removal from the low carbon effluents in intensive recirculating aquaculture systems (RAS). In this study, we tested three additional carbon sources, including biochar, dried Sphagnum sp. moss and industrial potato residues, to enhance the performance of woodchip bioreactors treating the low carbon RAS wastewater. We assessed nitrate (NO3−) removal and microbial community composition during a one-y…
Molecular diversity and growth features of Flavobacterium columnare strains isolated in Finland
ABSTRACT: Columnaris disease caused by Flavobacterium columnare is a problem in fish farmingworldwide. During the last 15 yr, outbreaks have started to emerge in Finland. Flavobacteriumcolumnare Type Strain NCIMB 2248 T and 30 Finnish F. columnare isolates were studied usinganalysis of 16S rDNA by restriction-fragment length polymorphism (16S RFLP), length heterogeneityanalysis of polymerase chain reaction (LH-PCR) products, automated ribosomal intergenic spaceranalysis (ARISA), and 16S rDNA sequence analysis. All isolates fell into RFLP Genomovar I and hadthe same length in the LH-PCR analysis. Based on ARISA, 8 genetically different strains wereselected for further analyses. The growth of…
Biofiltration of airborne VOCs with green wall systems : microbial and chemical dynamics
Botanical air filtration is a promising technology for reducing indoor air contaminants, but the underlying mechanisms need better understanding. Here, we made a set of chamber fumigation experiments of up to 16 weeks of duration, to study the filtration efficiencies for seven volatile organic compounds (VOCs; decane, toluene, 2‐ethylhexanol, α‐pinene, octane, benzene, and xylene) and to monitor microbial dynamics in simulated green wall systems. Biofiltration functioned on sub‐ppm VOC levels without concentration‐dependence. Airflow through the growth medium was needed for efficient removal of chemically diverse VOCs, and the use of optimized commercial growth medium further improved the e…
Successful aerobic bioremediation of groundwater contaminated with higher chlorinated phenols by indigenous degrader bacteria
The xenobiotic priority pollutant pentachlorophenol has been used as a timber preservative in a polychlorophenol bulk synthesis product containing also tetrachlorophenol and trichlorophenol. Highly soluble chlorophenol salts have leaked into groundwater, causing severe contamination of large aquifers. Natural attenuation of higher-chlorinated phenols (HCPs: pentachlorophenol + tetrachlorophenol) at historically polluted sites has been inefficient, but a 4-year full scale in situ biostimulation of a chlorophenol-contaminated aquifer by circulation and re-infiltration of aerated groundwater was remarkably successful: pentachlorophenol decreased from 400 μg L−1 to <1 μg L−1 and tetrachlorophen…
Methanotrophy induces nitrogen fixation during peatland development
Significance In peatlands, the external sources of nitrogen are mainly atmospheric, but the atmospheric nitrogen deposition alone cannot explain the long-term annual nitrogen accumulation rates to these ecosystems. Because of methodological problems, methane-induced fixation of atmospheric dinitrogen gas has been previously overlooked as an additional nitrogen input mechanism. We found that the activity of methane-oxidizing bacteria provides not only carbon but also nitrogen to peat mosses and, thus, contributes to carbon and nitrogen accumulation in peatlands, which store approximately one-third of the global soil carbon pool. Our results imply that nitrogen fixation in wetlands may be str…
Diversity of chlorophenol-degrading bacteria isolated from contaminated boreal groundwater
Chlorophenol-degrading bacteria from a long-term polluted groundwater aquifer were characterized. All isolates degraded 2,4,6-trichlorophenol and 2,3,4,6-tetrachlorophenol at concentrations detected in the contaminated groundwater (10 mg 1(-1)). Pentachlorophenol was degraded by three isolates when present alone. In two gram-positive isolates, 2,3,4,6-tetrachlorophenol was required as an inducer for the degradation of pentachlorophenol. The gram-positive isolates were sensitive to pentachlorophenol, with an IC50 value of 5 mg/l. Isolates belonging to the Cytophaga/Flexibacter/Bacteroides phylum had IC50 values of 25 and 63 mg/l. Isolates belonging to alpha-, beta- and gamma-Proteobacteria g…
Isolation and Characterization of Novosphingobium sp. Strain MT1, a Dominant Polychlorophenol-Degrading Strain in a Groundwater Bioremediation System
ABSTRACT A high-rate fluidized-bed bioreactor has been treating polychlorophenol-contaminated groundwater in southern Finland at 5 to 8°C for over 6 years. We examined the microbial diversity of the bioreactor using three 16S ribosomal DNA (rDNA)-based methods: denaturing gradient gel electrophoresis, length heterogeneity-PCR analysis, and restriction fragment length polymorphism analysis. The molecular study revealed that the process was dependent on a stable bacterial community with low species diversity. The dominant organism, Novosphingobium sp. strain MT1, was isolated and characterized. Novosphingobium sp. strain MT1 degraded the main contaminants of the groundwater, 2,4,6-trichloroph…
Effect of peracetic acid on levels of geosmin, 2-methylisoborneol, and their potential producers in a recirculating aquaculture system for rearing rainbow trout (Oncorhynchus mykiss)
Abstract In recirculating aquaculture systems (RAS)s, off-flavors and odors, mainly caused by geosmin (GSM) and 2-methylisoborneol (MIB), can accumulate in the flesh of fish from RAS water, reducing the profitability of production. In this study, peracetic acid (PAA) was applied in three application intervals to pump sumps of rainbow trout (Oncorhynchus mykiss) reared in RAS. Using a real-time polymerase chain reaction (qPCR), the potential off-flavor producers were quantified using geoA and MIB synthase genes. Streptomyces was identified as the major GSM producer, and biofilters showed the highest number of potential off-flavor producers. Concentrations of GSM and MIB were analyzed in the …
Green sulphur bacteria as a component of the photosynthetic plankton community in small dimictic humic lakes with an anoxic hypolimnion
High bacteriochlorophyll (BChl) concentrations in the anoxic water layers of some humic lakes have indicated that green sulphur bacteria (GSB) may be ecologically significant. The abundance and spatial distribution of GSB were therefore addressed in 13 small humic lakes using fragment analysis and sequencing of PCR-amplified 16S rRNA genes. GSB were detected from lakes where the photosynthetically active radiation was at least 1.1 µE m �2 s �1 at the oxic� anoxic boundary layer. In these lakes, 13 to 42% of the 16S rRNA gene sequences of the anoxic water column were assigned to GSB. The spatial distribution of GSB was tightly correlated with the spectrophotometrically measured BChl concentr…
Evidence of enhanced bacterial invasion during Diplostomum spathaceum infection in European grayling, Thymallus thymallus (L.)
Farmed grayling, Thymallus thymallus (L.), are susceptible to atypical Aeromonas salmonicida (aAS) infections. Interactions between bacteria and parasites were studied using grayling subjected to concomitant exposure to aAS bacteria and the digenean parasite Diplostomum spathaceum. Atypical AS was detected from fish by a combination of bacterial cultivation and polymerase chain reaction techniques. A detection level of 17 aAS cells per 100 mg intestine tissue sample was obtained. Concomitant bacterial exposure did not enhance the severity of grayling eye rupture and nuclear extrusion induced by D. spathaceum, but D. spathaceum invasion into grayling increased the proportion of fish carrying…
Effect of Pseudomonas sp. MT5 baths on Flavobacterium columnare infection of rainbow trout and on microbial diversity on fish skin and gills
Use of Pseudomonas sp. strain MT5 to prevent and treat Flavobacterium columnare infection was studied in 2 experiments with fingerling rainbow trout Oncorhynchus mykiss. In the first experiment, length heterogeneity analysis of PCR-amplified DNA fragments (LH-PCR) was used to assess the effect of antagonistic baths on the microbial diversity of healthy and experimentally infected fish. In the 148 samples studied, no difference was found between bathed and unbathed fish, and 3 fragment lengths were detected most frequently: 500 (in 75.7% of the samples), 523 (62.2%) and 517 bp (40.5%). The species contributing to these fragment sizes were Pseudomonas sp., Rhodococcus sp. and F. columnare, re…
CH4 oxidation in a boreal lake during the development of hypolimnetic hypoxia
AbstractFreshwater ecosystems represent a significant natural source of methane (CH4). CH4 produced through anaerobic decomposition of organic matter (OM) in lake sediment and water column can be either oxidized to carbon dioxide (CO2) by methanotrophic microbes or emitted to the atmosphere. While the role of CH4 oxidation as a CH4 sink is widely accepted, neither the magnitude nor the drivers behind CH4 oxidation are well constrained. In this study, we aimed to gain more specific insight into CH4 oxidation in the water column of a seasonally stratified, typical boreal lake, particularly under hypoxic conditions. We used 13CH4 incubations to determine the active CH4 oxidation sites and the …
A practical method for barcoding and size-trimming PCR templates for amplicon sequencing
Sample barcoding facilitates the analysis of tens or even hundreds of samples in a single next-generation sequencing (NGS) run, but more efficient methods are needed for high-throughput barcoding and size-trimming of long PCR products. Here we present a two-step PCR approach for barcoding followed by pool shearing, adapter ligation, and 5′ end selection for trimming sets of DNA templates of any size. Our new trimming method offers clear benefits for phylogenetic studies, since targeting exactly the same region maximizes the alignment and enables the use of operational taxonomic unit (OTU)-based algorithms.
Genetic diversity and phenotypic characterization of Iodobacter limnosediminis associated with skin lesions in freshwater fish
The relatively unknown genus Iodobacter sp. has been repeatedly isolated from skin ulcers and saprolegniosis on freshwater fish in Finland, especially farmed salmonids. Genetic characterization verified that all 23 bacterial isolates studied here belonged to the species Iodobacter limnosediminis, previously undescribed from the fish microbiota. Whole-genome pulsed-field gel electrophoresis revealed variability between the I. limnosediminis strains, suggesting that they were most likely of environmental origin. Two I. limnosediminis strains caused lesions in 27%–53% of brown trout (Salmo trutta) injected intramuscularly (p ≤ .05). The lesions represented moderate to severe tissue damage, but…
DNA recovery from Droplet Digital™ PCR emulsions using liquid nitrogen
Droplet microfluidics is a technology that enables the production and manipulation of small volumes. In biosciences, the most popular application of this technology is Droplet Digital™ PCR (ddPCR™), where parallel nanoliter-scale PCR assays are used to provide a high sensitivity and specificity for DNA detection. However, the recovery of PCR products for downstream applications such as sequencing can be challenging due to the droplets' stability. Here we compared five methods for disrupting the droplets to recover DNA. We found that rapid freezing in liquid nitrogen results in a clear phase separation and recovery of up to 70% of the DNA content. Liquid nitrogen freezing can thus offer a si…
Nicotinamide riboside improves muscle mitochondrial biogenesis, satellite cell differentiation, and gut microbiota in a twin study
Nicotinamide adenine dinucleotide (NAD + ) precursor nicotinamide riboside (NR) has emerged as a promising compound to improve obesity-associated mitochondrial dysfunction and metabolic syndrome in mice. However, most short-term clinical trials conducted so far have not reported positive outcomes. Therefore, we aimed to determine whether long-term NR supplementation boosts mitochondrial biogenesis and metabolic health in humans. Twenty body mass index (BMI)–discordant monozygotic twin pairs were supplemented with an escalating dose of NR (250 to 1000 mg/day) for 5 months. NR improved systemic NAD + metabolism, muscle mitochondrial number, myoblast differentiation, and gut microbiota compos…
Grazing intensity in subarctic tundra affects the temperature adaptation of soil microbial communities
Abstract Grazing by large ungulates, such as reindeer (Rangifer tarandus L.), in subarctic tundra exerts a considerable effect on the soil microclimate. Because of higher insulation by the aboveground vegetation in light versus heavily grazed areas, soil temperatures during the growing season are considerably higher under heavy grazing. Here, we hypothesized that these grazer-induced changes in soil microclimate affect the temperature sensitivity of soil microbial activity. To test this hypothesis, we conducted soil incubations at different temperatures (4 °C, 9 °C and 14 °C) for six weeks using soils from sites with contrasting long-term grazing intensities. Microbial respiration at low te…
Distinct and diverse anaerobic bacterial communities in boreal lakes dominated by candidate division OD1
Lakes have a central role in the carbon cycle of the boreal landscape. These systems typically stratify in summer and their hypolimnetic microbial communities influence burial of biogenic organic matter in sediments. The composition of bacterial communities in these suboxic habitats was studied by pyrosequencing of 16S rRNA amplicons from five lakes with variable dissolved organic carbon (DOC) concentrations. Bacterioplankton communities in the hypolimnetic waters were clearly different from the surface layer with candidate division OD1, Chlorobi and Bacteroidetes as dominant community members. Several operational taxonomic units (OTUs) affiliated with candidate division OD1 were abundant a…
Enrichment of Bacterioplankton Able to Utilize One-Carbon and Methylated Compounds in the Coastal Pacific Ocean
International audience; Understanding the temporal variations and succession of bacterial communities involved in the turnover of one-carbon and methylated compounds is necessary to better predict bacterial impacts on the marine carbon cycle and air-sea carbon fluxes. The ability of the local bacterioplankton community to exploit one-carbon and methylated compounds as main source of bioavailable carbon during a productive and less productive period was assessed through enrichment experiments. Surface seawater was amended with methanol and trimethylamine-N-oxide (TMAO), and bacterial abundance, production, oxygen consumption, as well as methanol turnover and growth rates of putative methylot…
Prebiotic Xylo-oligosaccharides Targeting Faecalibacterium prausnitzii Prevent High Fat Diet-induced Hepatic Steatosis in Rats
Understanding the importance of gut microbiota (GM) in non-alcoholic fatty liver disease (NAFLD) has raised the hope for therapeutic microbes. We have shown that high hepatic fat associated with low abundance of Faecalibacterium prausnitzii in humans and further, administration of F. prausnitzii prevented NAFLD in mice. Here, we aimed to target F. prausnitzii by prebiotic xylo-oligosaccharides (XOS) to treat NAFLD. First, the effect of XOS on F. prausnitzii growth was assessed in vitro. Then, XOS was supplemented or not with high (HFD) or low (LFD) fat-diet for 12-weeks in Wistar rats (n=10/group). XOS increased F. prausnitzii growth having only minor impact on the GM composition. When supp…
Vertical stratification patterns of methanotrophs and their genetic controllers in water columns of oxygen-stratified boreal lakes
ABSTRACT The vertical structuring of methanotrophic communities and its genetic controllers remain understudied in the water columns of oxygen-stratified lakes. Therefore, we used 16S rRNA gene sequencing to study the vertical stratification patterns of methanotrophs in two boreal lakes, Lake Kuivajärvi and Lake Lovojärvi. Furthermore, metagenomic analyses were performed to assess the genomic characteristics of methanotrophs in Lovojärvi and the previously studied Lake Alinen Mustajärvi. The methanotroph communities were vertically structured along the oxygen gradient. Alphaproteobacterial methanotrophs preferred oxic water layers, while Methylococcales methanotrophs, consisting of putative…
Sphingobium aromaticiconvertens sp. nov., a xenobiotic-compound-degrading bacterium from polluted river sediment.
A bacterial strain capable of degrading some monochlorinated dibenzofurans, designated RW16T, was isolated from aerobic River Elbe sediments. The strain was characterized based on 16S rRNA gene sequence analysis, DNA G+C content, physiological characteristics, polyamines, ubiquinone and polar lipid pattern and fatty acid composition. This analysis revealed that strain RW16T represents a novel species of the genus Sphingobium. The DNA G+C content of strain RW16T, 60.7 mol%, is the lowest yet reported for the genus. 16S rRNA gene sequence analysis placed strain RW16T as an outlier in the genus Sphingobium. The name Sphingobium aromaticiconvertens sp. nov. is proposed for this dibenzofuran-min…
Salinity affects nitrate removal and microbial composition of denitrifying woodchip bioreactors treating recirculating aquaculture system effluents
Abstract This study investigated the effect of salinity on microbial composition and denitrification capacity of woodchip bioreactors treating recirculating aquaculture system (RAS) effluents. Twelve laboratory-scale woodchip bioreactors were run in triplicates at 0, 15, 25, and 35 ppt salinities, and water chemistry was monitored every third day during the first 39 days of operation. Microbial communities of the woodchips bioreactors were analyzed at the start, after one week, and at the end of the trial. Woodchip bioreactors removed nitrate at all salinities tested. The highest NO3-N removal rate of 22.0 ± 6.9 g NO3-N/m3/d was obtained at 0 ppt, while 15.3 ± 4.9, 12.5 ± 5.4 and 11.8 ± 4.0…
Do shifts in life strategies explain microbial community responses to increasing nitrogen in tundra soil?
Subarctic tundra soils store large quantities of the global organic carbon (C) pool as the decomposition of plant litter and soil organic matter is limited by low temperatures and limiting nutrients. Mechanisms that drive organic matter decomposition are still poorly understood due to our limited knowledge of microbial communities and their responses to changing conditions. In subarctic tundra large grazers, in particular reindeer, exert a strong effect on vegetation and nutrient availability causing drastic nutrient pulses in the soils located along the migratory routes. Here we studied the effect of increased nitrogen (N) availability on microbial community structure and activities by lab…
Evidence for Natural Horizontal Transfer of the pcpB Gene in the Evolution of Polychlorophenol-Degrading Sphingomonads
ABSTRACT The chlorophenol degradation pathway in Sphingobium chlorophenolicum is initiated by the pcpB gene product, pentachlorophenol-4-monooxygenase. The distribution of the gene was studied in a phylogenetically diverse group of polychlorophenol-degrading bacteria isolated from contaminated groundwater in Kärkölä, Finland. All the sphingomonads isolated were shown to share pcpB gene homologs with 98.9 to 100% sequence identity. The gene product was expressed when the strains were induced by 2,3,4,6-tetrachlorophenol. A comparative analysis of the 16S rDNA and pcpB gene trees suggested that a recent horizontal transfer of the pcpB gene was involved in the evolution of the catabolic pat…
Eutrophication reduces the nutritional value of phytoplankton in boreal lakes
Eutrophication (as an increase in total phosphorus [TP]) increases harmful algal blooms and reduces the proportion of high-quality phytoplankton in seston and the content of ω-3 long-chain polyunsaturated fatty acids (eicosapentaenoic acid [EPA] and docosahexaenoic acid [DHA]) in fish. However, it is not well-known how eutrophication affects the overall nutritional value of phytoplankton. Therefore, we studied the impact of eutrophication on the production (as concentration; μg L−1) and content (μg mg C−1) of amino acids, EPA, DHA, and sterols, i.e., the nutritional value of phytoplankton in 107 boreal lakes. The lakes were categorized in seven TP concentration categories ranging from ultra…
Enhanced nitrogen removal of low carbon wastewater in denitrification bioreactors by utilizing industrial waste toward circular economy
Abstract Aquaculture needs practical solutions for nutrient removal to achieve sustainable fish production. Passive denitrifying bioreactors may provide an ecological, low-cost and low-maintenance approach for wastewater nitrogen removal. However, innovative organic materials are needed to enhance nitrate removal from the low carbon effluents in intensive recirculating aquaculture systems (RAS). In this study, we tested three additional carbon sources, including biochar, dried Sphagnum sp. moss and industrial potato residues, to enhance the performance of woodchip bioreactors treating the low carbon RAS wastewater. We assessed nitrate (NO3−) removal and microbial community composition durin…
Decomposition rate and biochemical fate of carbon from natural polymers and microplastics in boreal lakes
Microbial mineralization of organic compounds is essential for carbon recycling in food webs. Microbes can decompose terrestrial recalcitrant and semi-recalcitrant polymers such as lignin and cellulose, which are precursors for humus formation. In addition to naturally occurring recalcitrant substrates, microplastics have been found in various aquatic environments. However, microbial utilization of lignin, hemicellulose, and microplastics as carbon sources in freshwaters and their biochemical fate and mineralization rate in freshwaters is poorly understood. To fill this knowledge gap, we investigated the biochemical fate and mineralization rates of several natural and synthetic polymer-deri…
Comparing the Ecotoxicological Effects of Perfluorooctanoic Acid (PFOA) and Perfluorohexanoic Acid (PFHxA) on Freshwater Microbial Community
The ubiquitous presence of perfluorinated carboxylic acids (PFCAs) around the globe has attracted increasing attention, due to their persistency, bioaccumulation, and toxicity. Nevertheless, the ecotoxicological effects of the compounds on aquatic microorganisms has remained understudied. Hence, the present study focused on determining, and comparing, the effects of regulated long-chain PFCA, perfluorooctanoic acid (PFOA), and nonregulated short-chain PFCA, perfluorohexanoic acid (PFHxA), on the diversity, structure, microbial growth, and activity of a freshwater microbial community. In the experiment, lake water was incubated for a period of four weeks at three different concentrations of …
Stable isotope profiles of nitrogen gas indicate denitrification in oxygen-stratified humic lakes
Mid-summer N(2) profiles were analyzed from nine oxygen-stratified, humic-acid-rich lakes using a continuous flow isotope ratio mass spectrometer and a Gasbench II device. Sample preparation steps were performed under water to avoid air contamination. The instrument precision for the δ(15)N measurement was high (0.03‰), but for the whole sampling and analysis procedure the mean deviation between replicate samples was 0.13‰ for the δ(15)N measurements and 5.5% for the N(2) gas concentration analysis. The results show that the Gasbench peripheral was suitable for measurement of the (15)N natural abundance of dissolved nitrogen gas, with denitrification indicated by the oversaturation and slig…
Methanotrophs are core members of the diazotroph community in decaying Norway spruce logs
Dead wood is initially a nitrogen (N) poor substrate, where the N content increases with decay, partly due to biological N2 fixation, but the drivers of the N accumulation are poorly known. We quantified the rate of N2 fixation in decaying Norway spruce logs of different decay stages and studied the potential regulators of the N2-fixation activity. The average rate for acetylene reduction in the decaying wood was 7.5 nmol ethylene g−1d−1, which corresponds to 52.9 μg N kg−1d−1. The number of nifH copies (g−1 dry matter) was higher at the later decay stages, but no correlation between the copy number and the in vitro N2 fixation rate was found. All recovered nifH sequences were assigned to t…
Microbial Community Response on Wastewater Discharge in Boreal Lake Sediments
Despite high performance, municipal wastewater treatment plants (WWTPs) still discharge significant amounts of organic material and nitrogen and even microbes into the receiving water bodies, altering physico-chemical conditions and microbial functions. In this study, we examined how nitrified wastewater affects the microbiology of boreal lake sediments. Microbial community compositions were assessed with next generation sequencing of the 16S rRNA gene, and a more detailed view on nitrogen transformation processes was gained with qPCR targeting on functional genes (nirS, nirK, nosZI, nosZII, amoAarchaea, and amoAbacteria). In both of the two studied lake sites, the microbial community compo…
Effects of soybean meal based diet on growth performance, gut histopathology and intestinal microbiota of juvenile rainbow trout (Oncorhynchus mykiss)
Abstract Effects of soybean meal (SBM) based diet on growth performance, histology of the intestinal epithelium and on the gut microbiota of juvenile rainbow trout (Oncorhynchus mykiss) were investigated on two trials lasting for 8 weeks (Trial I) and 18 weeks (Trial II). The microbiological characterization was done both with conventional plating techniques, biochemical profiling and length heterogeneity analysis of PCR amplified 16S rDNA (LH-PCR). Typical histological changes were found after 18 weeks of SBM feeding (Trial II). Mean height of simple foldings were significantly higher within the group fed with fish meal (FM) based diet. These changes were not, however, reflected in the app…
Spatial and temporal changes in Actinobacterial dominance in experimental artificial groundwater recharge.
Abstract Artificial groundwater recharge (AGR) is used in the drinking water industry to supplement groundwater resources and to minimise the use of chemicals in water treatment. This study analysed the spatial and temporal changes of microbial communities in AGR using two test systems: a nutrient-amended fluidized-bed reactor (FBR) and a sand column. Structural changes in the feed lake water (Lake Roine), FBR, and sand column bacterial communities were determined by denaturing gradient gel electrophoresis (DGGE) and the length heterogeneity analysis of amplified 16S rRNA genes (LH-PCR). Two clone libraries were created to link the LH-PCR results to the dominant bacterial groups. The lake w…
Irradiation of the head reduces adult hippocampal neurogenesis and impairs spatial memory, but leaves overall health intact in rats.
Treatment of brain cancer, glioma, can cause cognitive impairment as a side‐effect, possibly because it disrupts the integrity of the hippocampus, a structure vital for normal memory. Radiotherapy is commonly used to treat glioma, but the effects of irradiation on the brain are still poorly understood, and other biological effects have not been extensively studied. Here we exposed healthy adult male rats to small and moderate‐dose irradiation of the head. We found no effect of irradiation on systemic inflammation, weight gain or gut microbiota diversity, although it increased the abundance of Bacteroidaceae family, namely Bacteroides genus in the gut microbiota. Irradiation had no effect on…
Bacterial community response to changes in a tri-trophic cascade during a whole-lake fish manipulation
Microbial communities play a key role in biogeochemical processes by degrading organic material and recycling nutrients, but can also be important food sources for upper trophic levels. Trophic cascades might modify microbial communities either directly via grazing or indirectly by inducing changes in other biotic or in abiotic factors (e.g., nutrients). We studied the effects of a tri-trophic cascade on microbial communities during a whole-lake manipulation in which European perch (Perca fluviatilis) were added to a naturally fishless lake divided experimentally into two basins. We measured environmental parameters (oxygen, temperature, and nutrients) and zooplankton biomass and studied th…
Cooperation between Different CRISPR-Cas Types Enables Adaptation in an RNA-Targeting System
CRISPR-Cas systems are immune systems that protect bacteria and archaea against their viruses, bacteriophages. Immunity is achieved through the acquisition of short DNA fragments from the viral invader’s genome.
Consistency of Targeted Metatranscriptomics and Morphological Characterization of Phytoplankton Communities
The composition of phytoplankton community is the basis for environmental monitoring and assessment of the ecological status of aquatic ecosystems. Community composition studies of phytoplankton have been based on time-consuming and expertise-demanding light microscopy analyses. Molecular methods have the potential to replace microscopy, but the high copy number variation of ribosomal genes and the lack of universal primers for simultaneous amplification of prokaryotic and eukaryotic genes complicate data interpretation. In this study, we used our previously developed directional primer-independent high-throughput sequencing (HTS) approach to analyze 16S and 18S rRNA community structures. C…
Subtercola boreus gen. nov., sp. nov. and Subtercola frigoramans sp. nov., two new psychrophilic actinobacteria isolated from boreal groundwater.
Psychrophilic actinobacterial isolates from permanently cold groundwater in Finland were characterized using a polyphasic approach. Growth on agar plates was observed at temperatures down to -2 degrees C, with an optimum at 15-17 degrees C, but no growth was observed at 30 degrees C. The peptidoglycan type was B2y and the characteristic diamino acid was diaminobutyric acid. The cell wall sugars of strain K265T were rhamnose, ribose, xylose and mannose and those of strain K300T were glucose, rhamnose and xylose. The polar lipids included phosphatidylglycerol, diphosphatidylglycerol, one unknown phospholipid and two glycolipids. The main whole-cell fatty acids were 12-methyltetradecanoic acid…
The effect of iron on the biodegradation of natural dissolved organic matter
Iron (Fe) may alter the biodegradation of dissolved organic matter (DOM), by interacting with DOM, phosphorus (P), and microbes. We isolated DOM and a bacterial community from boreal lake water and examined bacterial growth on DOM in laboratory experiments. Fe was introduced either together with DOM (DOM-Fe) or into bacterial suspension, which led to the formation of insoluble Fe precipitates on bacterial surfaces (Fe coating). In the latter case, the density of planktonic bacteria was an order of magnitude lower than that in the corresponding treatment without introduced Fe. The association of Fe with DOM decreased bacterial growth, respiration, and growth efficiency compared with DOM alon…
Microbial Adaptation to Boreal Saturated Subsurface: Implications in Bioremediation of Polychlorophenols
Saturated subsurface environments pose challenges to the intrinsic microbiology. Prevailing environmental conditions (temperature, pH, bioavailability of substrates and nutrients) affect microbial biodegradation activity, which is often favored by certain redox conditions. Microbial adaptation in each redox environment proceeds by selection and enrichment of indigenous bacteria, evolution of novel catabolic pathways and horizontal gene transfer (Wilson et al. 1985; van der Meer et al. 1998; Tiirola et al. 2002b). Formation of biofilms enables microbial retainment, co-operation among microorganisms and enhanced gene transfer among organisms (Singh et al. 2006). Chlorophenols are toxic and pe…
Microbial diversity in a thermophilic aerobic biofilm process: analysis by length heterogeneity PCR (LH-PCR).
A two-stage pilot-scale thermophilic aerobic suspended carrier biofilm process (SCBP) was set up for the on-site treatment of pulp and paper mill whitewater lining. The microbial diversity in this process was analyzed by length heterogeneity analysis of PCR-amplified 16S ribosomal DNA. The primer pair selected for PCR amplification was first evaluated by a computational analysis of fragment lengths in ten main phylogenetical eubacterial groups. The fragment contained the first third of the 16S rRNA gene, which was shown to vary naturally between 465 and 563 bp in length. The length heterogeneity analysis of polymerase chain reaction (LH-PCR) profile of the biomass attached to carrier elemen…
Nitrate removal microbiology in woodchip bioreactors : a case-study with full-scale bioreactors treating aquaculture effluents
Woodchip bioreactors are viable low-cost nitrate (NO3−) removal applications for treating agricultural and aquaculture discharges. The active microbial biofilms growing on woodchips are conducting nitrogen (N) removal, reducing NO3− while oxidizing the carbon (C) from woodchips. However, bioreactor age, and changes in the operating conditions or in the microbial community might affect the NO3− removal as well as potentially promote nitrous oxide (N2O) production through either incomplete denitrification or dissimilatory NO3− reduction to ammonium (DNRA). Here, we combined stable isotope approach, amplicon sequencing, and captured metagenomics for studying the potential NO3− removal rates, a…
Sediment diffusion method improves wastewater nitrogen removal in the receiving lake sediments
Sediment microbes have a great potential to transform reactive N to harmless N2, thus decreasing wastewater nitrogen load into aquatic ecosystems. Here, we examined if spatial allocation of the wastewater discharge by a specially constructed sediment diffuser pipe system enhanced the microbial nitrate reduction processes. Full-scale experiments were set on two Finnish lake sites, Keuruu and Petäjävesi, and effects on the nitrate removal processes were studied using the stable isotope pairing technique. All nitrate reduction rates followed nitrate concentrations, being highest at the wastewater-influenced sampling points. Complete denitrification with N2 as an end-product was the main nitrat…
Comparison of epifluorescence microscopy and flow cytometry in counting freshwater picophytoplankton
The smaller the phytoplankton, the greater effort is required to distinguish individual cells by optics-based methods. Flow cytometry is widely applied in marine picophytoplankton research, but in freshwater research its role has remained minor. We compared epifluorescence microscopy and flow cytometry in assessing the composition, abundance and cell sizes of autofluorescent picophytoplankton in epilimnia of 46 Finnish lakes. Phycocyaninrich picocyanobacteria were the most dominant. The two methods yielded comparable total picophytoplankton abundances, but the determination of cell sizes, and thus total biomasses, were on average an order of magnitude higher in the microscopy results. Howev…
Microbial diversity along a gradient in peatlands treating mining-affected waters.
Peatlands are used for the purification of mining-affected waters in Northern Finland. In Northern climate, microorganisms in treatment peatlands (TPs) are affected by long and cold winters, but studies about those microorganisms are scarce. Thus, the bacterial, archaeal and fungal communities along gradients of mine water influence in two TPs were investigated. The TPs receive waters rich in contaminants, including arsenic (As), sulfate (SO42-) and nitrate (NO3-). Microbial diversity was high in both TPs, and microbial community composition differed between the studied TPs. Bacterial communities were dominated by Proteobacteria, Actinobacteria, Chloroflexi and Acidobacteria, archaeal commu…
New insight to the role of microbes in the methane exchange in trees: evidence from metagenomic sequencing
Methane (CH4) exchange in tree stems and canopies and the processes involved are among the least understood components of the global CH4 cycle. Recent studies have focused on quantifying tree stems as sources of CH4 and understanding abiotic CH4 emissions in plant canopies, with the role of microbial in situ CH4 formation receiving less attention. Moreover, despite initial reports revealing CH4 consumption, studies have not adequately evaluated the potential of microbial CH4 oxidation within trees. In this paper, we discuss the current level of understanding on these processes. Further, we demonstrate the potential of novel metagenomic tools in revealing the involvement of microbes in the C…
Spatial and temporal variation in denitrification and in the denitrifier community in a boreal lake
We investigated the spatial and temporal variation in denitrification rates (isotopepairing technique, IPT) and in the denitrifier community (examination of gene nirK by denaturinggradient gel electrophoresis [DGGE] of microbial DNA) in the sediments of a boreal, clear-water, eutrophic lake using samples collected from shallow littoral, deep littoral and shallow profundal sediments during early summer, mid-summer, autumn and winter. The measured denitrification rates (44 to 613 μmol N m−2 d−1) are among the lowest ever reported from lacustrine sediments. Denitri fi - cation rates varied both spatially and temporally, being highest in the profundal zone during midsummer and in the littoral z…
Vertical diversity of bacteria in an oxygen-stratified humic lake, evaluated using DNA and phospholipid analyses
Microbes play a particularly important role in the food web in lakes with high dissolved organic carbon content. The bacterial community of a polyhumic lake, Mekkojarvi, was studied using DNA techniques and phospholipid fatty acid (PLFA) analysis during the mid-summer period of water column strati- fication. According to the 16S rRNA gene clone libraries and length heterogeneity analysis (LH-PCR), heterotrophic bacteria dominated only in the oxic epilimnion, in which various Actinobacteria (mostly cluster acI-B) and Betaproteobacteria (especially Poly- nucleobacter subcluster PnecC) were common. Se- quences assigned to heterotrophic, methylotrophic, photoautotrophic, and chemoautotrophic ge…
Treatment of columnaris disease of rainbow trout: low pH and salt as possible tools?
The impact of salt and low pH on columnaris disease of fish was studied. Survival of Flavobacterium columnare after exposure to either 4% NaCl (pH 7.2) or pH 5.0, pH 4.86 or pH 4.6 for 15 min or 1 h was studied in vitro. All conditions significantly reduced the numbers of viable bacterial cells. The effects of salt (4 and 2%) and acidic baths (pH 4.6) were studied in 2 experiments in vivo with rainbow trout Oncorhynchus mykiss infected with F. columnare. Both salt and acidic baths failed to prevent fish mortality; the overall mortality reached 100% in all groups. However, according to survival analysis, the mortality rate was lower in fish treated with 4% salt baths compared to a control gr…
Cryptogams signify key transitions of bacteria and fungi in Arctic sand dune succession.
•Primary succession models focus on aboveground vascular plants. However, the prevalence of mosses and lichens, i.e. cryptogams, suggests they play a role in soil successions. Here, we explore whether effects of cryptogams on belowground microbes can facilitate progressive shifts in sand dune succession. •We linked aboveground vegetation, belowground bacterial and fungal communities, and soil chemical properties in six successional stages in Arctic inland sand dunes: bare sand, grass, moss, lichen, ericoid heath and mountain birch forest. •Compared to the bare sand and grass stages, microbial biomass and the proportion of fungi increased in the moss stage, and later stage microbial groups a…
Probing the Gelation Synergies and Anti-Escherichia coli Activity of Fmoc-Phenylalanine/Graphene Oxide Hybrid Hydrogel
The N-fluorenyl-9-methyloxycarbonyl (Fmoc)-protected amino acids have shown high antimicrobial application potential, among which the phenylalanine derivative (Fmoc-F) is the most well-known representative. However, the activity spectrum of Fmoc-F is restricted to Gram-positive bacteria only. The demand for efficient antimicrobial materials expanded research into graphene and its derivatives, although the reported results are somewhat controversial. Herein, we combined graphene oxide (GO) flakes with Fmoc-F amino acid to form Fmoc-F/GO hybrid hydrogel for the first time. We studied the synergistic effect of each component on gelation and assessed the material’s bactericidal activity on Gram…
Resistant ammonia-oxidizing archaea endure, but adapting ammonia-oxidizing bacteria thrive in boreal lake sediments receiving nutrient-rich effluents
Climate change along with anthropogenic activities changes biogeochemical conditions in lake ecosystems, modifying the sediment microbial communities. Wastewater effluents introduce nutrients and organic material but also novel microbes to lake ecosystems, simulating forthcoming increases in catchment loadings. In this work, we first used 16s rRNA gene sequencing to study how the overall sediment microbial community responds to wastewater in six boreal lakes. To examine forthcoming changes in the lake biogeochemistry, we focused on the ammonia-oxidizing archaea (AOA) and bacteria (AOB), and examined their functional and compositional community response to wastewater. Although we found the l…
Abundant and diverse arsenic‐metabolizing microorganisms in peatlands treating arsenic‐contaminated mining wastewaters
Mining operations produce large quantities of wastewater. At a mine site in Northern Finland, two natural peatlands are used for the treatment of mining‐influenced waters with high concentrations of sulfate and potentially toxic arsenic (As). In the present study, As removal and the involved microbial processes in those treatment peatlands (TPs) were assessed. Arsenic‐metabolizing microorganisms were abundant in peat soil from both TPs (up to 108 cells gdw‐1), with arsenate respirers being about 100 times more abundant than arsenite oxidizers. In uninhibited microcosm incubations, supplemented arsenite was oxidized under oxic conditions and supplemented arsenate was reduced under anoxic con…
The effect of peracetic acid on microbial community, water quality, nitrification and rainbow trout (Oncorhynchus mykiss) performance in recirculating aquaculture systems
Microbial biofilters control water quality and enable the overall function of recirculation aquaculture systems (RAS). Changes in environmental conditions can affect the abundance and interactions of the diverse microbial populations of the biofilter, affecting nitrification of harmful ammonium and thus fish health. Here, we examined the effect of different application frequencies (0, 1, 2 and 4 times per week) of a common disinfectant, peracetic acid (PAA, applied 1.1 mg l−1 twice per day), on biofilter microbial communities, focusing especially on nitrifying microbial groups and using a high throughput sequencing of 16S rRNA gene and quantitative PCR (qPCR). In addition, we measured biofi…
Denitrifying microbial communities along a boreal stream with varying land-use
Streams have an important role in regulating nitrogen (N) transportation from terrestrial ecosystems to downstream waters. Here, we examined how catchment land-use affects potential denitrification rates and the function and composition of denitrifier communities in boreal stream sediments, using stable isotope incubations and qPCR and 454-pyrosequencing targeted on nirS, nirK and nosZ genes. Although land-use influenced the water chemistry as higher nitrite + nitrate (NO x − ) concentration at the agriculture-affected sampling point, sediment organic matter content was found to be the key factor in regulating potential denitrification rates. However, the abundance as well as the diversity …
Bacterial and phytoplankton responses to nutrient amendments in a boreal lake differ according to season and to taxonomic resolution
Nutrient limitation and resource competition in bacterial and phytoplankton communities may appear different when considering different levels of taxonomic resolution. Nutrient amendment experiments conducted in a boreal lake on three occasions during one open water season revealed complex responses in overall bacterioplankton and phytoplankton abundance and biovolume. In general, bacteria were dominant in spring, while phytoplankton was clearly the predominant group in autumn. Seasonal differences in the community composition of bacteria and phytoplankton were mainly related to changes in observed taxa, while the differences across nutrient treatments within an experiment were due to chang…
Supplementary figure: DNA recovery from Droplet Digital PCR emulsions using Liquid Nitrogen
Supplementary figure 1 – Applicability of LN2 method to break ddPCR emulsion. ddPCR reactions were prepared with ddPCR™ Supermix for Probes (no dUTP) and Droplet Generation Oil for Probes (three tubes on the left) or ddPCR™ Supermix for EvaGreen® and Droplet Generation Oil for EvaGreen® (three tubes on the right). The tubes (A) and (D) show the emulsion before using LN2 method, and tubes (B), (C), (E) and (F) shows the oil and water layers after breaking the emulsion.
Supplementary table: DNA recovery from Droplet Digital PCR emulsions using Liquid Nitrogen
Supplementary table - Raw data from experiments studying DNA recovery from droplets using chemical and physical procedures.
Appendix A. Potential methane oxidation rates detailed by Sphagnum species, habitat, water level, and methane concentration.
Potential methane oxidation rates detailed by Sphagnum species, habitat, water level, and methane concentration.
Original data for article: Comparison of epifluorescence microscopy and flow cytometry in counting freshwater picophytoplankton
The dataset is divided into four subfolders: 1) "SEM experiment data" contains Scanning Electron Microscopy data, epifluorescence microscopy data and flow cytometry data of cultured Synechococcus, Chroococcus and Snowella 2) "raw data" contains epifluorescence microscopy and flow cytometry data of picophytoplankton from Finnish lakes. This has two sub folders "flow cytometry raw" and "microscopy raw" 3) "flow cytometry calibration data" contains data for cell size calibration with latex beads and volumetric calibration for the flow cytometer 4) "processed flow and microscopy data" contains excel workbooks for the figures shown in the manuscipt