Resilience of aerobic methanotrophs in soils; spotlight on the methane sink under agriculture.

Aerobic methanotrophs are a specialized microbial group, catalyzing the oxidation of methane. Disturbance-induced loss of methanotroph diversity/abundance, thus results in the loss of this biological methane sink. Here, we synthesized and conceptualized the resilience of the methanotrophs to sporadic, recurring, and compounded disturbances in soils. The methanotrophs showed remarkable resilience to sporadic disturbances, recovering in activity and population size. However, activity was severely compromised when disturbance persisted or reoccurred at increasing frequency, and was significantly impaired following change in land use. Next, we consolidated the impact of agricultural practices after land conversion on the soil methane sink. The effects of key interventions (tillage, organic matter input, and cover cropping) where much knowledge has been gathered were considered. Pairwise comparisons of these interventions to nontreated agricultural soils indicate that the agriculture-induced impact on the methane sink depends on the cropping system, which can be associated to the physiology of the methanotrophs. The impact of agriculture is more evident in upland soils, where the methanotrophs play a more prominent role than the methanogens in modulating overall methane flux. Although resilient to sporadic disturbances, the methanotrophs are vulnerable to compounded disturbances induced by anthropogenic activities, significantly affecting the methane sink function.

Specialized biopolymers: versatile tools for microbial resilience.

Bacteria produce a wide range of specialized biopolymers that can be classified into polysaccharides, polyamides, and polyesters and are considered to fulfill storage functions. In this review, we highlight recent developments in the field linking metabolism of biopolymers to stress and signaling physiology of the producers and demonstrating that biopolymers contribute to bacterial stress resistance and shape structure and composition of microenvironments. While specialized biopolymers are currently the focus of much attention in biotechnology as innovative and biodegradable materials, our understanding about the regulation and functions of these valuable compounds for the producers, microbial communities, and our environment is still very limited. Addressing open questions about signals, mechanisms, and functions in the area of biopolymers harbors potential for exciting discoveries with high relevance for biotechnology and fundamental research.

Setting new standards: Multiphasic analysis of microplastic mineralization by fungi.

Plastic materials provide numerous benefits. However, properties such as durability and resistance to degradation that make plastic attractive for variable applications likewise foster accumulation in the environment. Fragmentation of plastics leads to the formation of potentially hazardous microplastic, of which a considerable amount derives from polystyrene. Here, we investigated the biodegradation of polystyrene by the tropical sooty mold fungus Capnodium coffeae in different experimental setups. Growth of C. coffeae was stimulated significantly when cultured in presence of plastic polymers rather than in its absence. Stable isotope tracing using 13C-enriched polystyrene particles combined with cavity ring-down spectroscopy showed that the fungus mineralized polystyrene traces. However, phospholipid fatty acid stable isotope probing indicated only marginal assimilation of polystyrene-13C by C. coffeae in liquid cultures. NMR spectroscopic analysis of residual styrene contents prior to and after incubation revealed negligible changes in concentration. Thus, this study suggests a plastiphilic life style of C. coffeae despite minor usage of plastic as a carbon source and the general capability of sooty mold fungi to stimulate polystyrene mineralization, and proposes new standards to identify and unambiguously demonstrate plastic degrading capabilities of microbes.

Biodegradable polymers boost reproduction in the earthworm Eisenia fetida.

Microplastic contamination in soil has become a global environmental threat as it adversely affects terrestrial organisms like earthworms as well as soil properties. Especially biodegradable polymers have recently been used as an alternative to conventional polymer types, although their impact remains poorly understood. Thus, we studied the effect of conventional (polystyrene: PS, polyethylene terephthalate: PET, polypropylene: PP) versus aliphatic polyesters classified as biodegradable polymers (poly-(l-lactide): PLLA, polycaprolactone: PCL) on the earthworm Eisenia fetida and soil properties (pH and cation exchange capacity). We addressed direct effects on the weight gain and reproductive success of E. fetida, and indirect effects, like changes in the gut microbial composition as well as the production of short-chain fatty acids by the gut microbiota. Earthworms were exposed for eight weeks in an artificial soil amended with two environmentally relevant concentrations (1 % and 2.5 % (w/w)) of the different microplastic types. PLLA and PCL boosted the number of cocoons produced by 135 % and 54 %, respectively. Additionally, exposure to these two polymers increased number of hatched juveniles, changed gut microbial beta-diversity, and increased the production of the short chain fatty acid lactate compared to the control treatments. Interestingly, we also found a positive effect of PP on the earthworm's bodyweight and reproductive success. The interaction of microplastic and earthworms decreased soil pH by about 1.5 units in the presence of PLLA and PCL. No polymer effect on the cation exchange capacity of soil was found. In general, neither the presence of conventional nor biodegradable polymers had any adverse effects on any of the studied endpoints. Our results suggest that the effects of microplastic highly depend on the polymer type, and that the degradation of biodegradable polymers might be enhanced in the gut of earthworms, which implies that they may use biodegradable polymers as a potential carbon source.

Interkingdom interaction: the soil isopod Porcellio scaber stimulates the methane-driven bacterial and fungal interaction.

Porcellio scaber (woodlice) are (sub-)surface-dwelling isopods, widely recognized as "soil bioengineers", modifying the edaphic properties of their habitat, and affecting carbon and nitrogen mineralization that leads to greenhouse gas emissions. Yet, the impact of soil isopods on methane-cycling processes remains unknown. Using P. scaber as a model macroinvertebrate in a microcosm study, we determined how the isopod influences methane uptake and the associated interaction network in an agricultural soil. Stable isotope probing (SIP) with 13C-methane was combined to a co-occurrence network analysis to directly link activity to the methane-oxidizing community (bacteria and fungus) involved in the trophic interaction. Compared to microcosms without the isopod, P. scaber significantly induced methane uptake, associated to a more complex bacteria-bacteria and bacteria-fungi interaction, and modified the soil nutritional status. Interestingly, 13C was transferred via the methanotrophs into the fungi, concomitant to significantly higher fungal abundance in the P. scaber-impacted soil, indicating that the fungal community utilized methane-derived substrates in the food web along with bacteria. Taken together, results showed the relevance of P. scaber in modulating methanotrophic activity with implications for bacteria-fungus interaction.

Mitomycin C-induced effects on aerobic methanotrophs in a landfill cover soil; implications of a viral shunt?

A viral shunt can occur when phages going through a lytic cycle, including lysogenic phages triggered by inducing agents (e.g. mitomycin C), results in host lysis and the release of cell constituents and virions. The impact of a viral shunt on the carbon, including methane cycle in soil systems is poorly understood. Here, we determined the effects of mitomycin C on the aerobic methanotrophs in a landfill cover soil. To an extent, our results support a mitomycin C-induced viral shunt, as indicated by the significantly higher viral-like particle (VLP) counts relative to bacteria, elevated nutrient concentrations (ammonium, succinate), and initially impaired microbial activities (methane uptake and microbial respiration) after mitomycin C addition. The trend in microbial activities at <2 days largely corresponded to the expression of the pmoA and 16S rRNA genes. Thereafter (>11 days), the active bacterial community composition significantly diverged in the mitomycin C-supplemented incubations, suggesting the differential impact of mitomycin C on the bacterial community. Collectively, we provide insight on the effects of mitomycin C, and potentially a viral shunt, on the bacteria in the soil environment.

Microplastic ingestion affects hydrogen production and microbiomes in the gut of the terrestrial isopod Porcellio scaber.

Microplastic (MP) is an environmental burden and enters food webs via ingestion by macrofauna, including isopods (Porcellio scaber) in terrestrial ecosystems. Isopods represent ubiquitously abundant, ecologically important detritivores. However, MP-polymer specific effects on the host and its gut microbiota are unknown. We tested the hypothesis that biodegradable (polylactic acid [PLA]) and non-biodegradable (polyethylene terephthalate [PET]; polystyrene [PS]) MPs have contrasting effects on P. scaber mediated by changes of the gut microbiota. The isopod fitness after an 8-week MP-exposure was generally unaffected, although the isopods showed avoidance behaviour to PS-food. MP-polymer specific effects on gut microbes were detected, including a stimulation of microbial activity by PLA compared with MP-free controls. PLA stimulated hydrogen emission from isopod guts, while PET and PS were inhibitory. We roughly estimated 107 kg year-1 hydrogen emitted from the isopods globally and identified their guts as anoxic, significant mobile sources of reductant for soil microbes despite the absence of classical obligate anaerobes, likely due to Enterobacteriaceae-related fermentation activities that were stimulated by lactate generated during PLA-degradation. The findings suggest negative effects of PET and PS on gut fermentation, modulation of important isopod hydrogen emissions by MP pollution and the potential of MP to affect terrestrial food webs.

Intermediate Levels of Predation and Nutrient Enrichment Enhance the Activity of Ibuprofen-Degrading Bacteria.

Water is the most indispensable natural resource; yet, organic pollution of freshwater sources is widespread. In recent years, there has been increasing concern over the vast array of emerging organic contaminants (EOCs) in the effluent of wastewater treatment plants (WWTPs). Several of these EOCs are degraded within the pore space of riverbeds by active microbial consortia. However, the mechanisms behind this ecosystem service are largely unknown. Here, we report how phosphate concentration and predator-prey interactions drive the capacity of bacteria to process a model EOC (ibuprofen). The presence of phosphate had a significant positive effect on the population growth rate of an ibuprofen-degrading strain. Thus, when phosphate was present, ibuprofen removal efficiency increased. Moreover, low and medium levels of predation, by a ciliated protozoan, stimulated bacterial population growth. This unimodal effect of predation was lost under high phosphate concentration, resulting in the flattening of the relationships between predator density and population growth of ibuprofen degraders. Our results suggest that moderate nutrient and predation levels promote the growth rate of bacterial degraders and, consequently, the self-purifying capability of the system. These findings enhance our understanding of the mechanisms by which riverbed communities drive the processing of EOCs.

Consumption of N2O by Flavobacterium azooxidireducens sp. nov. Isolated from Decomposing Leaf Litter of Phragmites australis (Cav.).

Microorganisms acting as sinks for the greenhouse gas nitrous oxide (N2O) are gaining increasing attention in the development of strategies to control N2O emissions. Non-denitrifying N2O reducers are of particular interest because they can provide a real sink without contributing to N2O release. The bacterial strain under investigation (IGB 4-14T), isolated in a mesocosm experiment to study the litter decomposition of Phragmites australis (Cav.), is such an organism. It carries only a nos gene cluster with the sec-dependent Clade II nosZ and is able to consume significant amounts of N2O under anoxic conditions. However, consumption activity is considerably affected by the O2 level. The reduction of N2O was not associated with cell growth, suggesting that no energy is conserved by anaerobic respiration. Therefore, the N2O consumption of strain IGB 4-14T rather serves as an electron sink for metabolism to sustain viability during transient anoxia and/or to detoxify high N2O concentrations. Phylogenetic analysis of 16S rRNA gene similarity revealed that the strain belongs to the genus Flavobacterium. It shares a high similarity in the nos gene cluster composition and the amino acid similarity of the nosZ gene with various type strains of the genus. However, phylogenomic analysis and comparison of overall genome relatedness indices clearly demonstrated a novel species status of strain IGB 4-14T, with Flavobacterium lacus being the most closely related species. Various phenotypic differences supported a demarcation from this species. Based on these results, we proposed a novel species Flavobacterium azooxidireducens sp. nov. (type strain IGB 4-14T = LMG 29709T = DSM 103580T).

A globally relevant stock of soil nitrogen in the Yedoma permafrost domain.

Nitrogen regulates multiple aspects of the permafrost climate feedback, including plant growth, organic matter decomposition, and the production of the potent greenhouse gas nitrous oxide. Despite its importance, current estimates of permafrost nitrogen are highly uncertain. Here, we compiled a dataset of >2000 samples to quantify nitrogen stocks in the Yedoma domain, a region with organic-rich permafrost that contains ~25% of all permafrost carbon. We estimate that the Yedoma domain contains 41.2 gigatons of nitrogen down to ~20 metre for the deepest unit, which increases the previous estimate for the entire permafrost zone by ~46%. Approximately 90% of this nitrogen (37 gigatons) is stored in permafrost and therefore currently immobile and frozen. Here, we show that of this amount, ¾ is stored >3 metre depth, but if partially mobilised by thaw, this large nitrogen pool could have continental-scale consequences for soil and aquatic biogeochemistry and global-scale consequences for the permafrost feedback.

Microplastic polymer properties as deterministic factors driving terrestrial plastisphere microbiome assembly and succession in the field.

Environmental microplastic (MP) is ubiquitous in aquatic and terrestrial ecosystems providing artificial habitats for microbes. Mechanisms of MP colonization, MP polymer impacts, and effects on soil microbiomes are largely unknown in terrestrial systems. Therefore, we experimentally tested the hypothesis that MP polymer type is an important deterministic factor affecting MP community assembly by incubating common MP polymer types in situ in landfill soil for 14 months. 16S rRNA gene amplicon sequencing indicated that MP polymers have specific impacts on plastisphere microbiomes, which are subsets of the soil microbiome. Chloroflexota, Gammaproteobacteria, certain Nitrososphaerota, and Nanoarchaeota explained differences among MP polymers and time points. Plastisphere microbial community composition derived from different MP diverged over time and was enriched in potential pathogens. PICRUSt predictions of pathway abundances and quantitative PCR of functional marker genes indicated that MP polymers exerted an ambivalent effect on genetic potentials of biogeochemical cycles. Overall, the data indicate that (i) polymer type as deterministic factor rather than stochastic factors drives plastisphere community assembly, (ii) MP impacts greenhouse gas metabolism, xenobiotic degradation and pathogen distribution, and (iii) MP serves as an ideal model system for studying fundamental questions in microbial ecology such as community assembly mechanisms in terrestrial environments.

Microbial iron reduction compensates for phosphorus limitation in paddy soils.

Limitation of rice growth by low phosphorus (P) availability is a widespread problem in tropical and subtropical soils because of the high content of iron (Fe) (oxyhydr)oxides. Ferric iron-bound P (Fe(III)-P) can serve as a P source in paddies after Fe(III) reduction to Fe(II) and corresponding H2PO4- release. However, the relevance of reductive dissolution of Fe(III)-P for plant and microbial P uptake is still an open question. To quantify this, 32P-labeled ferrihydrite (30.8 mg P kg-1) was added to paddy soil mesocosms with rice to trace the P uptake by microorganisms and plants after Fe(III) reduction. Nearly 2% of 32P was recovered in rice plants, contributing 12% of the total P content in rice shoots and roots after 33 days. In contrast, 32P recovery in microbial biomass decreased from 0.5% to 0.08% of 32P between 10 and 33 days after rice transplantation. Microbial biomass carbon (MBC) and dissolved organic C content decreased from day 10 to 33 by 8-54% and 68-77%, respectively, suggesting that the microbial-mediated Fe(III) reduction was C-limited. The much faster decrease of MBC in rooted (by 54%) vs. bulk soil (8-36%) reflects very fast microbial turnover in the rice rhizosphere (high C and oxygen inputs) resulting in the mineralization of the microbial necromass. In conclusion, Fe(III)-P can serve as small but a relevant P source for rice production and could partly compensate plant P demand. Therefore, the P fertilization strategies should consider the P mobilization from Fe (oxyhydr)oxides in flooded paddy soils during rice growth. An increase in C availability for microorganisms in the rhizosphere intensifies P mobilization, which is especially critical at early stages of rice growth.

The methane-driven interaction network in terrestrial methane hotspots.

BACKGROUND: Biological interaction affects diverse facets of microbial life by modulating the activity, diversity, abundance, and composition of microbial communities. Aerobic methane oxidation is a community function, with emergent community traits arising from the interaction of the methane-oxidizers (methanotrophs) and non-methanotrophs. Yet little is known of the spatial and temporal organization of these interaction networks in naturally-occurring complex communities. We hypothesized that the assembled bacterial community of the interaction network in methane hotspots would converge, driven by high substrate availability that favors specific methanotrophs, and in turn influences the recruitment of non-methanotrophs. These environments would also share more co-occurring than site-specific taxa. RESULTS: We applied stable isotope probing (SIP) using 13C-CH4 coupled to a co-occurrence network analysis to probe trophic interactions in widespread methane-emitting environments, and over time. Network analysis revealed predominantly unique co-occurring taxa from different environments, indicating distinctly co-evolved communities more strongly influenced by other parameters than high methane availability. Also, results showed a narrower network topology range over time than between environments. Co-occurrence pattern points to Chthoniobacter as a relevant yet-unrecognized interacting partner particularly of the gammaproteobacterial methanotrophs, deserving future attention. In almost all instances, the networks derived from the 13C-CH4 incubation exhibited a less connected and complex topology than the networks derived from the unlabelledC-CH4 incubations, likely attributable to the exclusion of the inactive microbial population and spurious connections; DNA-based networks (without SIP) may thus overestimate the methane-dependent network complexity. CONCLUSION: We demonstrated that site-specific environmental parameters more strongly shaped the co-occurrence of bacterial taxa than substrate availability. Given that members of the interactome without the capacity to oxidize methane can exert interaction-induced effects on community function, understanding the co-occurrence pattern of the methane-driven interaction network is key to elucidating community function, which goes beyond relating activity to community composition, abundances, and diversity. More generally, we provide a methodological strategy that substantiates the ecological linkages between potentially interacting microorganisms with broad applications to elucidate the role of microbial interaction in community function.

Genomic evidence for two pathways of formaldehyde oxidation and denitrification capabilities of the species Paracoccus methylovorus sp. nov.

Three strains (H4-D09T, S2-D11 and S9-F39) of a member of the genus Paracoccus attributed to a novel species were isolated from topsoil of temperate grasslands. The genome sequence of the type strain H4-D09T exhibited a complete set of genes required for denitrification as well as methylotrophy. The genome of H4-D09T included genes for two alternative pathways of formaldehyde oxidation. Besides the genes for the canonical glutathione (GSH)-dependent formaldehyde oxidation pathway, all genes for the tetrahydrofolate-formaldehyde oxidation pathway were identified. The strain has the potential to utilize methanol and/or methylamine as a single carbon source as evidenced by the presence of methanol dehydrogenase (mxaFI) and methylamine dehydrogenase (mau) genes. Apart from dissimilatory denitrification genes (narA, nirS, norBC and nosZ), genes for assimilatory nitrate (nasA) and nitrite reductases (nirBD) were also identified. The results of phylogenetic analysis based on 16S rRNA genes coupled with riboprinting revealed that all three strains represented the same species of genus Paracoccus. Core genome phylogeny of the type strain H4-D09T indicated that Paracoccus thiocyanatus and Paracoccus denitrificans are the closest phylogenetic neighbours. The average nucleotide index (ANI) and digital DNA-DNA hybridization (dDDH) with the closest phylogenetic neighbours revealed genetic differences at the species level, which were further substantiated by differences in several physiological characteristics. The major respiratory quinone is Q-10, and the predominant cellular fatty acids are C18 : 1ω7c, C19 : 0cyclo ω7c, and C16 : 0, which correspond to those detected in other members of the genus. The polar lipid profile consists of a diphosphatidylglycerol (DPG), phosphatidylethanolamine (PE), phosphatidylglycerol (PG), phosphatidylcholine (PC), aminolipid (AL), glycolipid (GL) and an unidentified lipid (L).On the basis of our results, we concluded that the investigated isolates represent a novel species of the genus Paracoccus, for which the name Paracoccus methylovorus sp. nov. (type strain H4-D09T=LMG 31941T= DSM 111585T) is proposed.

Aerobic Methanotrophy and Co-occurrence Networks of a Tropical Rainforest and Oil Palm Plantations in Malaysia.

Oil palm (OP) plantations are gradually replacing tropical rainforest in Malaysia, one of the largest palm oil producers globally. Conversion of lands to OP plantations has been associated with compositional shifts of the microbial community, with consequences on the greenhouse gas (GHG) emissions. While the impact of the change in land use has recently been investigated for microorganisms involved in N2O emission, the response of the aerobic methanotrophs to OP agriculture remains to be determined. Here, we monitored the bacterial community composition, focusing on the aerobic methanotrophs, in OP agricultural soils since 2012, 2006, and 1993, as well as in a tropical rainforest, in 2019 and 2020. High-affinity methane uptake was confirmed, showing significantly lower rates in the OP plantations than in the tropical rainforest, but values increased with continuous OP agriculture. The bacterial, including the methanotrophic community composition, was modified with ongoing OP agriculture. The methanotrophic community composition was predominantly composed of unclassified methanotrophs, with the canonical (Methylocystis) and putative methanotrophs thought to catalyze high-affinity methane oxidation present at higher relative abundance in the oldest OP plantation. Results suggest that the methanotrophic community was relatively more stable within each site, exhibiting less temporal variations than the total bacterial community. Uncharacteristically, a 16S rRNA gene-based co-occurrence network analysis revealed a more complex and connected community in the OP agricultural soil, which may influence the resilience of the bacterial community to disturbances. Overall, we provide a first insight into the ecology and role of the aerobic methanotrophs as a methane sink in OP agricultural soils.

Biodegradation of metoprolol in oxic and anoxic hyporheic zone sediments: unexpected effects on microbial communities.

Metoprolol is widely used as a beta-blocker and considered an emerging contaminant of environmental concern due to pseudo persistence in wastewater effluents that poses a potential ecotoxicological threat to aquatic ecosystems. Microbial removal of metoprolol in the redox-delineated hyporheic zone (HZ) was investigated using streambed sediments supplemented with 15 or 150 µM metoprolol in a laboratory microcosm incubation under oxic and anoxic conditions. Metoprolol disappeared from the aqueous phase under oxic and anoxic conditions within 65 and 72 days, respectively. Metoprolol was refed twice after initial depletion resulting in accelerated disappearance under both conditions. Metoprolol disappearance was marginal in sterile control microcosms with autoclaved sediment. Metoprolol was transformed mainly to metoprolol acid in oxic microcosms, while metoprolol acid and α-hydroxymetoprolol were formed in anoxic microcosms. Transformation products were transient and disappeared within 30 days under both conditions. Effects of metoprolol on the HZ bacterial community were evaluated using DNA- and RNA-based time-resolved amplicon Illumina MiSeq sequencing targeting the 16S rRNA gene and 16S rRNA, respectively, and were prominent on 16S rRNA rather than 16S rRNA gene level suggesting moderate metoprolol-induced activity-level changes. A positive impact of metoprolol on Sphingomonadaceae and Enterobacteriaceae under oxic and anoxic conditions, respectively, was observed. Nitrifiers were impaired by metoprolol under oxic and anoxic conditions. Collectively, our findings revealed high metoprolol biodegradation potentials in the hyporheic zone under contrasting redox conditions associated with changes in the active microbial communities, thus contributing to the attenuation of micropollutants. KEY POINTS: • High biotic oxic and anoxic metoprolol degradation potentials in the hyporheic zone. • Key metoprolol-associated taxa included Sphingomonadaceae, Enterobacteraceae, and Promicromonosporaceae. • Negative impact of metoprolol on nitrifiers.

Microplastics accumulate fungal pathogens in terrestrial ecosystems.

Microplastic (MP) is a pervasive pollutant in nature that is colonised by diverse groups of microbes, including potentially pathogenic species. Fungi have been largely neglected in this context, despite their affinity for plastics and their impact as pathogens. To unravel the role of MP as a carrier of fungal pathogens in terrestrial ecosystems and the immediate human environment, epiplastic mycobiomes from municipal plastic waste from Kenya were deciphered using ITS metabarcoding as well as a comprehensive meta-analysis, and visualised via scanning electron as well as confocal laser scanning microscopy. Metagenomic and microscopic findings provided complementary evidence that the terrestrial plastisphere is a suitable ecological niche for a variety of fungal organisms, including important animal and plant pathogens, which formed the plastisphere core mycobiome. We show that MPs serve as selective artificial microhabitats that not only attract distinct fungal communities, but also accumulate certain opportunistic human pathogens, such as cryptococcal and Phoma-like species. Therefore, MP must be regarded a persistent reservoir and potential vector for fungal pathogens in soil environments. Given the increasing amount of plastic waste in terrestrial ecosystems worldwide, this interrelation may have severe consequences for the trans-kingdom and multi-organismal epidemiology of fungal infections on a global scale.

Transformation of organic micropollutants along hyporheic flow in bedforms of river-simulating flumes.

Urban streams receive increasing loads of organic micropollutants from treated wastewaters. A comprehensive understanding of the in-stream fate of micropollutants is thus of high interest for water quality management. Bedforms induce pumping effects considerably contributing to whole stream hyporheic exchange and are hotspots of biogeochemical turnover processes. However, little is known about the transformation of micropollutants in such structures. In the present study, we set up recirculating flumes to examine the transformation of a set of micropollutants along single flowpaths in two triangular bedforms. We sampled porewater from four locations in the bedforms over 78 days and analysed the resulting concentration curves using the results of a hydrodynamic model in combination with a reactive transport model accounting for advection, dispersion, first-order removal and retardation. The four porewater sampling locations were positioned on individual flowpaths with median solute travel times ranging from 11.5 to 43.3 h as shown in a hydrodynamic model previously. Highest stability was estimated for hydrochlorothiazide on all flowpaths. Lowest detectable half-lives were estimated for sotalol (0.7 h) and sitagliptin (0.2 h) along the shortest flowpath. Also, venlafaxine, acesulfame, bezafibrate, irbesartan, valsartan, ibuprofen and naproxen displayed lower half-lives at shorter flowpaths in the first bedform. However, the behavior of many compounds in the second bedform deviated from expectations, where particularly transformation products, e.g. valsartan acid, showed high concentrations. Flowpath-specific behavior as observed for metformin or flume-specific behavior as observed for metoprolol acid, for instance, was attributed to potential small-scale or flume-scale heterogeneity of microbial community compositions, respectively. The results of the study indicate that the shallow hyporheic flow field and the small-scale heterogeneity of the microbial community are major controlling factors for the transformation of relevant micropollutants in river sediments.