Microplastics (<5 mm) have been documented in environmental samples on a global scale. While these pollutants may enter aquatic environments via wastewater treatment facilities, the abundance of ...microplastics in these matrices has not been investigated. Although efficient methods for the analysis of microplastics in sediment samples and marine organisms have been published, no methods have been developed for detecting these pollutants within organic-rich wastewater samples. In addition, there is no standardized method for analyzing microplastics isolated from environmental samples. In many cases, part of the identification protocol relies on visual selection before analysis, which is open to bias. In order to address this, a new method for the analysis of microplastics in wastewater was developed. A pretreatment step using 30% hydrogen peroxide (H2O2) was employed to remove biogenic material, and focal plane array (FPA)-based reflectance micro-Fourier-transform (FT-IR) imaging was shown to successfully image and identify different microplastic types (polyethylene, polypropylene, nylon-6, polyvinyl chloride, polystyrene). Microplastic-spiked wastewater samples were used to validate the methodology, resulting in a robust protocol which was nonselective and reproducible (the overall success identification rate was 98.33%). The use of FPA-based micro-FT-IR spectroscopy also provides a considerable reduction in analysis time compared with previous methods, since samples that could take several days to be mapped using a single-element detector can now be imaged in less than 9 h (circular filter with a diameter of 47 mm). This method for identifying and quantifying microplastics in wastewater is likely to provide an essential tool for further research into the pathways by which microplastics enter the environment.
The marine benthic nitrogen cycle is affected by both the presence and activity of macrofauna and the diversity of N-cycling microbes. However, integrated research simultaneously investigating ...macrofauna, microbes and N-cycling is lacking. We investigated spatio-temporal patterns in microbial community composition and diversity, macrofaunal abundance and their sediment reworking activity, and N-cycling in seven subtidal stations in the Southern North Sea.
Our results indicated that bacteria (total and β-AOB) showed more spatio-temporal variation than archaea (total and AOA) as sedimentation of organic matter and the subsequent changes in the environment had a stronger impact on their community composition and diversity indices in our study area. However, spatio-temporal patterns of total bacterial and β-AOB communities were different and related to the availability of ammonium for the autotrophic β-AOB. Highest bacterial richness and diversity were observed in June at the timing of the phytoplankton bloom deposition, while richness of β-AOB as well as AOA peaked in September. Total archaeal community showed no temporal variation in diversity indices.
Distance based linear models revealed that, independent from the effect of grain size and the quality and quantity of sediment organic matter, nitrification and N-mineralization were affected by respectively the diversity of metabolically active β-AOB and AOA, and the total bacteria, near the sediment-water interface. Separate models demonstrated a significant and independent effect of macrofaunal activities on community composition and richness of total bacteria, and diversity indices of metabolically active AOA. Diversity of β-AOB was significantly affected by macrofaunal abundance. Our results support the link between microbial biodiversity and ecosystem functioning in marine sediments, and provided broad correlative support for the hypothesis that this relationship is modulated by macrofaunal activity. We hypothesized that the latter effect can be explained by their bioturbating and bio-irrigating activities, increasing the spatial complexity of the biogeochemical environment.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
For many organisms, there is agreement on the specific genomic region used for developing barcode markers. With nematodes, however, it has been found that the COI region designated for most animals ...lacks the taxonomic coverage (ability to amplify a diverse group of taxa) required of a metabarcoding marker. For that reason, studies on metabarcoding of nematodes thus far have utilized primarily regions within the highly conserved 18S ribosomal DNA. Two popular markers within this region are the ones flanked by the primer pairs NF1-18Sr2b and SSUF04-SSUR22. The NF1-18Sr2b primer pair, especially, has been critiqued as not being specific enough for nematodes leading to suggestions for other candidate markers while the SSUF04-SSUR22 region has hardly been tested on soil nematodes. The current study aimed to evaluate these two markers against other alternative ones within the 28S rDNA and the COI region for their suitability for nematode metabarcoding. The results showed that the NF1-18Sr2b marker could offer wide coverage and good resolution for characterizing soil nematodes. Sufficient availability of reference sequences for this region was found to be a significant factor that resulted in this marker outperforming the other markers, particularly the 18S-based SSUFO4-SSUR22 marker. None of the other tested regions compared with this marker in terms of the proportion of the taxa recovered. The COI-based marker had the lowest number of taxa recovered, and this was due to the poor performance of its primers and the insufficient number of reference sequences in public databases. In summary, this study highlights how dependent the success of metabarcoding is on the availability of a good reference sequence collection for the marker of choice as well as its taxonomic coverage.
While the presence of microplastics has been reported in aquatic habitats across the globe, the pathways through which they enter the environment are still poorly understood. Studies investigating ...the fate of microplastics in wastewater are gaining attention but are still scarce, despite the urgent need to understand the role of wastewater treatment plants (WWTP) as point sources of aquatic microplastic pollution. A likely reason for the limited number of WWTP-associated studies is that working with a biogenic organic matter (BOM)-rich sample matrix like wastewater is challenging. Here, we investigated the presence of microplastics (MP) throughout several stages of a wastewater treatment plant (WWTP) at multiple depths, employing Fenton’s reagent and focal plane array-based reflectance micro-Fourier-transform infrared spectroscopic (FPA-based reflectance micro-FTIR) imaging, a protocol that allows the automated detection and identification of microplastics in complex samples with high organic matter content, without the need for previous visual sorting, or reducing considerably the thickness of the sample, or the use of IR-transparent transmission windows. It was found that the number of microplastic fragments detected at downstream stages of the WWTP notably decreased following the primary settlement stage, with primary settlement stage samples responsible for 76.9% of total microplastics detected. Despite the marked reduction in the number of microplastic particles following the primary settlement stage, an average total of 1.5 MP L-1 were identified in the final effluent of the WWTP.
Currently, little is known about the impact of silver nanoparticles (AgNPs) on ecologically important microorganisms such as ammonia‐oxidizing bacteria (AOB). We performed a multi‐analytical approach ...to demonstrate the effects of uncapped nanosilver (uAgNP), capped nanosilver (cAgNP) and Ag₂SO₄ on the activities of the AOB: Nitrosomonas europaea, Nitrosospira multiformis and Nitrosococcus oceani, and the growth of Escherichia coli and Bacillus subtilis as model bacterial systems in relation to AgNP type and concentration. All Ag treatments caused significant inhibition to the nitrification potential rates (NPRs) of Nitrosomonas europaea (decreased from 34 to < 16.7 μM NH₄ ⁺ oxidized day⁻¹), Nitrosospira multiformis (decreased from 46 to < 24.8 μM NH₄ ⁺ oxidized day⁻¹) and Nitrosococcus oceani (decreased from 26 to < 18.4 μM NH₄ ⁺ oxidized day⁻¹). Escherichia coli‐Ag interactions revealed that the percentage of damaged E. coli cells was 45% greater with Ag₂SO₄, 39% with cAgNPs and 33% with uAgNPs compared with controls. Generally, the inhibitory effect on AOB NPRs and E. coli/B. subtilis growth was in the following order Ag₂SO₄ > cAgNP > uAgNP. In conclusion, AgNPs (especially cAgNPs) and Ag₂SO₄ adversely affected AOB activities and thus have the potential to severely impact key microbially driven processes such as nitrification in the environment.
The phylum Oomycota comprises important tree pathogens like Phytophthora quercina, involved in central European oak decline, and Phytophthora cinnamomi shown to affect holm oaks among many other ...hosts. Despite the importance to study the distribution, dispersal and niche partitioning of this phylum, metabarcoding surveys, and studies considering environmental factors that could explain oomycete community patterns are still rare. We investigated oomycetes in the rhizosphere of evergreen oaks in a Spanish oak woodland using metabarcoding based on Illumina sequencing of the taxonomic marker cytochrome c oxidase subunit II (cox2). We developed an approach amplifying a 333 bp long fragment using the forward primer Hud‐F (Mycologia, 2000) and a reverse primer found using DegePrime (Applied and Environmental Microbiology, 2014). Factors reflecting topo‐edaphic conditions and tree health were linked to oomycete community patterns. The majority of detected OTUs belonged to the Peronosporales. Most taxa were relatives of the Pythiaceae, but relatives of the Peronosporaceae and members of the Saprolegniales were also found. The most abundant OTUs were related to Globisporangium irregulare and P. cinnamomi, both displaying strong site‐specific patterns. Oomycete communities were strongly correlated with the environmental factors: altitude, crown foliation, slope and soil skeleton and soil nitrogen. Our findings illustrate the significance of small scale variation in habitat conditions for the distribution of oomycetes and highlight the importance to study oomycete communities in relation to such ecological patterns.
The phylum Oomycota comprises important tree pathogens, but knowledge on environmental factors that could explain their community patterns is scarce. To advance our understanding, oomycetes in the oak rhizosphere were studied using metabarcoding of the taxonomic marker cytochrome c oxidase subunit II and linked to biotic and abiotic variables. Oomycete communities were strongly correlated with the environmental factors altitude, crown foliation, slope and soil skeleton and soil nitrogen.
The composition of bacterial communities associated with four diatom species was monitored during isolation and cultivation of algal cells. Strong shifts in the associated communities, linked with an ...increase in the numbers of phylotypes belonging to members of the Gammaproteobacteria, were observed during cultivation.
Rapid and reliable tools for detection and identification of plant parasitic nematodes are needed to prevent the introduction and spread of quarantine nematodes. A fast and simple DNA extraction ...method for target nematodes in nematode suspensions obtained from soil samples and a new quantitative real-time PCR method (qPCR) for the specific detection, identification and potential quantification of M. enterolobii were tested in an inter-laboratory comparison (ring test) to allow for a thorough evaluation of these molecular diagnostic tools. A test performance study involving seven laboratories was conducted to validate the developed protocols and to identify possible difficulties when implemented by diagnostic laboratories or national reference centers. Validation included test performance in terms of accuracy, analytical specificity, analytical sensitivity, repeatability, and reproducibility as defined by European Plant Protection Organization (EPPO) standard PM7/98. All positive and negative results for detection, identification and specificity were consistent between different laboratories despite different equipment used. Accuracy of real-time PCR was 100 % because test results and accepted reference values were in agreement. Analytical sensitivity results also matched between laboratories independent of the equipment used. The smallest amount of target DNA tested, two second-stage juveniles of M. enterolobii in a background of 500 non-target nematodes, was reliably detected by all labs. In addition, the repeatability and reproducibility of test results between laboratories was 100 %, even at the limit of detection. Thus, the inter-laboratory comparison showed the robustness of the developed methods and confirmed the in-house validation data.
The characterization of specific subsets of soil microbiota in the rhizosphere and endosphere has led to the recognition of plant species-specific microbiomes. Most attention has been given to ...microbial prokaryotes and fungi. Only recently was convincing evidence for the existence of plant species-specific protist microbiomes presented. Although protists are expected to shape the composition of bacterial and fungal communities and, thereby, directly impact plant health, a lack of cultures of these important plant-symbiotic protists has hampered their experimental exploration. To facilitate empirical plant microbiome research, we sampled Arabidopsis thaliana, established 79 cultures covering nearly all major groups of plant-symbiotic Cercozoa (protists), and have made these publicly available. We discuss our findings and propose potential roles that these protists may have in structuring the plant microbiome.
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