Jaak Truu

Microbial biomass, activity and community composition in constructed wetlands

The aim of the current article is to give an overview about microbial communities and their functioning but also about factors affecting microbial activity in the three most common types (surface flow and two types of sub-surface flow) of constructed wetlands. The paper reviews the community composition and structural diversity of the microbial biomass, analyzing different aspects of microbial activity with respect to wastewater properties, specific wetland type, and environmental parameters. A brief introduction about the application of different novel molecular techniques for the assessment of microbial communities in constructed wetlands is also given. Microbially mediated processes in constructed wetlands are mainly dependent on hydraulic conditions, wastewater properties, including substrate and nutrient quality and availability, filter material or soil type, plants, and different environmental factors. Microbial biomass is within similar ranges in both horizontal and vertical subsurface flow and surface flow constructed wetlands. Stratification of the biomass but also a stratified structural pattern of the bacterial community can be seen in subsurface flow systems. Microbial biomass C/N ratio is higher in horizontal flow systems compared to vertical flow systems, indicating the structural differences in microbial communities between those two constructed wetland types. The total activity of the microbial community is in the same range, but heterotrophic growth is higher in the subsurface (vertical flow) system compared to the surface flow systems. Available species-specific data about microbial communities in different types of wetlands is scarce and therefore it is impossible make any general conclusions about the dynamics of microbial community structure in wetlands, its relationship to removal processes and operational parameters.

PlutoF—a Web Based Workbench for Ecological and Taxonomic Research, with an Online Implementation for Fungal ITS Sequences

DNA sequences accumulating in the International Nucleotide Sequence Databases (INSD) form a rich source of information for taxonomic and ecological meta-analyses. However, these databases include many erroneous entries, and the data itself is poorly annotated with metadata, making it difficult to target and extract entries of interest with any degree of precision. Here we describe the web-based workbench PlutoF, which is designed to bridge the gap between the needs of contemporary research in biology and the existing software resources and databases. Built on a relational database, PlutoF allows remote-access rapid submission, retrieval, and analysis of study, specimen, and sequence data in INSD as well as for private datasets though web-based thin clients. In contrast to INSD, PlutoF supports internationally standardized terminology to allow very specific annotation and linking of interacting specimens and species. The sequence analysis module is optimized for identification and analysis of environmental ITS sequences of fungi, but it can be modified to operate on any genetic marker and group of organisms. The workbench is available at http://plutof.ut.ee.

Variation of microbiological parameters within planted soil filter for domestic wastewater treatment.

Abstract Microbial community structure was assessed in a horizontal subsurface flow planted sand filter treating domestic wastewater with molecular and culture-based methods. The diversity and spatial distribution of the microbial community was investigated using a PCR–DGGE (eubacterial and archaeal primers, ammonia-oxidizing bacteria, and ammonium monooxygenase specific primers), and spread plate and MPN counts. Significant differences were found in the spatial distribution of the microbial community structure. Data analysis revealed that different components of the microbial community possessed different spatial distribution patterns within the filter bed and depending on community type, relationships with soil chemical, and microbiological parameters varied. The most important spatial pattern in microbial community structure within the constructed wetland was related to the depth gradient, followed by differences between inflow and outflow. A comparison of a number of heterotrophic bacteria between inlet and outlet pipes as well as between two sampling depths showed no significant differences. In addition, the variation of the abundance of ammonia-oxidizing bacteria demonstrated no clear spatial pattern.

Dynamics of antibiotic resistance genes and their relationships with system treatment efficiency in a horizontal subsurface flow constructed wetland

Municipal wastewater treatment is one of the pathways by which antibiotic resistance genes from anthropogenic sources are introduced into natural ecosystems. This study examined the abundance and proportion dynamics of seven antibiotic resistance genes in the wetland media biofilm and in the influent and effluent of parallel horizontal subsurface flow mesocosm cells of a newly established hybrid constructed wetland treating municipal wastewater. The targeted genes (tetA, tetB, tetM, ermB, sul1, ampC, and qnrS) encode resistance to major antibiotic classes such as tetracyclines, macrolides, sulfonamides, penicillins, and fluoroquinolones, respectively. All targeted antibiotic resistance genes were detectable in the tested mesocosm environments, with the tetA, sul1, and qnrS genes being the most abundant in the mesocosm effluents. After initial fluctuation in the microbial community, target gene abundances and proportions stabilized in the wetland media biofilm. The abundance of 16S rRNA and antibiotic resistance genes, and the proportion of antibiotic resistance genes in the microbial community, were reduced during the wastewater treatment by the constructed wetland. The concentration of antibiotic resistance genes in the system effluent was similar to conventional wastewater treatment facilities; however, the mesocosms reduced sulfonamide resistance encoding sul1 concentrations more effectively than some traditional wastewater treatment options. The concentrations of antibiotic resistance genes in the wetland media biofilm and in effluent were affected by system operation parameters, especially time and temperature. The results also revealed a relationship between antibiotic resistance genes abundance and the removal efficiencies of NO2-N, NH4-N, and organic matter. Correlation analysis between the abundance of individual antibiotic resistance genes in the mesocosms influent, effluent and wetland media biofilm indicated that depending on antibiotic resistance gene type the microbes carrying these genes interact differently with microbial communities already present on the wetland media.

Microbial characteristics and nitrogen transformation in planted soil filter for domestic wastewater treatment.

Abstract We studied an experimental horizontal subsurface-flow planted sand filter in Kodijärve, Estonia. We measured the microbial biomass, nitrogen immobilization, potential nitrification, soil respiration, multiple carbon source utilization patterns of the microbial consortia of the soil samples, the carbon, nitrogen, and phosphorus content of the soil samples, the water quality and physicochemical indicators in water sampling wells as well as emissions of CO2, N2, NO2, and CH4 from the two beds (the dry bed and the wet bed) in the wetland. The potential nitrification of the upper layer of the dry bed could not be attributed primarily to autotrophic nitrification, or the nitrifying bacteria in this layer could be facultative heterotrophs, whereas autotrophic nitrification is predominant in the upper layer of the wet bed. It also was found that changing aeration conditions in the lower layer of the dry bed have resulted in a lower diversity of the microbial community and led to a relative depletion of easily degradable soil carbon resources.

Dynamic changes in the structure of microbial communities in Baltic Sea coastal seawater microcosms modified by crude oil, shale oil or diesel fuel

The coastal waters of the Baltic Sea are constantly threatened by oil spills, due to the extensive transportation of oil products across the sea. To characterise the hydrocarbon-degrading bacterial community of this marine area, microcosm experiments on diesel fuel, crude oil and shale oil were performed. Analysis of these microcosms, using alkane monooxygenase (alkB) and 16S rRNA marker genes in PCR-DGGE experiments, demonstrated that substrate type and concentration strongly influence species composition and the occurrence of alkB genes in respective oil degrading bacterial communities. Gammaproteobacteria (particularly the genus Pseudomonas) and Alphaproteobacteria were dominant in all microcosms treated with oils. All alkB genes carried by bacterial isolates (40 strains), and 8 of the 11 major DGGE bands from the microcosms, had more than 95% sequence identity with the alkB genes of Pseudomonas fluorescens. However, the closest relatives of the majority of sequences (54 sequences from 79) of the alkB gene library from initially collected seawater DNA were Actinobacteria. alkB gene expression, induced by hexadecane, was recorded in isolated bacterial strains. Thus, complementary culture dependent and independent methods provided a more accurate picture about the complex seawater microbial communities of the Baltic Sea.

Comparison of API 20NE and Biolog GN identification systems assessed by techniques of multivariate analyses

The increasing use of commercial multitest systems for identification of environmental bacteria creates the problem of how to compare the identification results obtained from different systems. The limited use of species designations in such comparisons is caused by low usage of environmental bacteria in the development of commercial identification schemes. Two multivariate statistical methods, the Mantels test and the co-inertia analysis, were applied to analyze data derived from the Biolog GN and the API 20NE systems of identification for 50 environmental bacterial strains. We found these two methods to be useful for revealing the relationship between the two sets of numerical taxonomic traits. Both of these methods showed that the distances according to the Biolog GN results between the studied strains were related to those derived from the API 20NE results, despite the differences in the test sets of the two systems. In addition, the co-inertia analysis allowed us to visualise the relationships between classifications of strains derived from the two identification systems and, simultaneously, to estimate the contribution of particular tests to the differentiation of bacterial strains.

Distribution of benthic diatoms in relation to environmental variables in lowland streams

The composition (% relative abundance) of diatom assemblages from soft bottom sediments was studied at 75 sites situated in 46 rivers, brooks, and ditches in the islands of Hiiumaa and Saaremaa and in the lowland of West Estonia. Although the total number of recorded taxa was 205, the studied diatom assemblages consisted of 54, 55, 48, and 50 constant species in the drainage basin of Moonsund, the Gulf of Riga, Hiiumaa, and Saaremaa, respectively. The habitats of the dominating taxa were heterogenous and the most widespread species were Achnanthidium minutissimum, Martyana martyi, Meridion circulare, Cocconeis placentula, Planothidium lanceolatum, and Amphora pediculus. The Shannon–Weaver diversity (H′) index varied from 2.09 to 4.63. Multivariate analyses were used to identify the environmental variables governing the composition and structure of the benthic diatom assemblage. Principal Component Analysis (PCA) and Redundancy Analysis (RDA), based on 56 most abundant taxa, indicated differences in the composition and structure of diatom assemblages between different drainage basins as well as between the upper and lower stream courses. In the headwaters there prevailed small epipsammic diatoms (Martyana, Planothidium, Staurosira, Staurosirella). Different motile epipelic species from the genera Amphora, Navicula, Nitzschia, etc. were distributed abundantly in the lower courses of the streams. There was a positive correlation between order of the stream site and trophic level of water (R=0.35; p<0.05). Along a river system, the increasing order of the stream was accompanied by higher trophic level of water.

Survival and catabolic performance of introduced Pseudomonas strains during phytoremediation and bioaugmentation field experiment.

A long-term field experiment was carried out to estimate the efficiency of bioaugmentation in combination with phytoremediation for oil shale chemical industry solid waste dump area remediation. Soil samples for microbiological and chemical analysis were collected during 3 years after bacterial biomass application. Microbial communities in soil samples were analysed using both culture-based and molecular methods. The survival of the introduced bacterial strains was confirmed by cultivation-based Box-PCR genomic fingerprints and denaturing gradient gel electrophoresis fingerprinting of the 16S rRNA and lmPH genes. The introduced bacterial strains as well as corresponding catabolic genes were recovered several years after biomass application, predominantly from the rhizosphere of birches. Soil samples from bioaugmented plots showed an elevated potential for degradation of phenolic compounds even 40 months after treatment. Based on our results we can conclude that the introduced Pseudomonas strains both survived, and their metabolic traits have persisted at the contaminated site over a long period of time.

Complementary seminovaginal microbiome in couples

The genital tract microbiome is tightly associated with reproductive health. Although many research studies have been performed on the vaginal microbiome, current knowledge of the male microbiome is scarce, and parallel studies examining couples are extremely rare. In this work, we aimed to compare seminal and vaginal microbiomes in couples and to assess the influence of sexual intercourse on vaginal microbiome. The study included 23 couples. Microbiomes of semen and vaginal fluid (pre- and post-intercourse) were profiled using Illumina HiSeq2000 sequencing of the V6 region of 16S rRNA gene. Seminal communities were significantly more diverse, but with lower total bacterial concentrations than those of the vagina. Gardnerella vaginalis was predominant in half of the women whose partners had significant leukocytospermia, but only in one of 17 women who had a partner without leukocytospermia. There was significant decrease in the relative abundance of Lactobacillus crispatus after intercourse, and high concordance between semen and vaginal samples. Our data support the hypothesis that semen and vaginal microbiomes are in association, inasmuch as the predominance of G. vaginalis in female partners was significantly related to inflammation in male genital tracts.