Dynamics of antibiotic resistance markers and Escherichia coli invasion in riverine heterotrophic biofilms facing increasing heat and flow stagnation.

As motivation to address environmental dissemination of antimicrobial resistance (AMR) is mounting, there is a need to characterize mechanisms by which AMR can propagate under environmental conditions. Here we investigated the effect of temperature and stagnation on the persistence of wastewater-associated antibiotic resistance markers in riverine biofilms and the invasion success of genetically-tagged Escherichia coli. Biofilms grown on glass slides incubated in-situ downstream of a wastewater treatment plant effluent discharge point were transferred to laboratory-scale flumes fed with filtered river water under potentially stressful temperature and flow conditions: recirculation flow at 20 °C, stagnation at 20 °C, and stagnation at 30 °C. After 14 days, quantitative PCR and amplicon sequencing were used to quantify bacteria, biofilms diversity, resistance markers (sul1, sul2, ermB, tetW, tetM, tetB, blaCTX-M-1, intI1) and E. coli. Resistance markers significantly decreased over time regardless of the treatment applied. Although invading E. coli were initially able to colonize the biofilms, its abundance subsequently declined. Stagnation was associated with a shift in biofilm taxonomic composition, but there was no apparent effect of flow conditions or the simulated river-pool warming (30 °C) on AMR persistence or invasion success of E. coli. Results however indicated that antibiotic resistance markers in the riverine biofilms decreased under the experimental conditions in the absence of exposure to external inputs of antibiotics and AMR.

Hydrological variations shape diversity and functional responses of streambed microbes.

Microbiota inhabiting the intermittent streambeds mediates several in-stream processes that are essential for ecosystem function. Reduced stream discharge caused by the strengthened intermittency and increased duration of the dry phase is a spreading global response to changes in climate. Here, the impacts of a 5-month desiccation, one-week rewetting and punctual storms, which interrupted the dry period, were examined. The genomic composition of total (DNA) and active (RNA) diversity, and the community level physiological profiles (CLPP) were considered as proxies for functional diversity to describe both prokaryotes and eukaryotes inhabiting the surface and hyporheic streambeds. Comparisons between the genomic and potential functional responses helped to understand how and whether the microbial diversity was sensitive to the environmental conditions and resource acquisition, such as water stress and extracellular enzyme activities, respectively. RNA expression showed the strongest relationship with the environmental conditions and resource acquisition, being more responsive to changing conditions compared to DNA diversity, especially in the case of prokaryotes. The DNA results presumably reflected the legacy of the treatments because inactive, dormant, or dead cells were included, suggesting a slow microbial biomass turnover or responses of the microbial communities to changes mainly through physiological acclimation. On the other hand, microbial functional diversity was largely explained by resources acquisition, such as metrics of extracellular enzymes, and appeared vulnerable to the hydrological changes and duration of desiccation. The data highlight the need to improve the functional assessment of stream ecosystems with the application of complementary metrics to better describe the streambed microbial dynamics under dry-rewet stress.

Distinct responses from bacterial, archaeal and fungal streambed communities to severe hydrological disturbances.

Stream microbes that occur in the Mediterranean Basin have been shown to possess heightened sensitivity to intensified water stress attributed to climate change. Here, we investigate the effects of long-term drought (150 days), storms and rewetting (7 days) on the diversity and composition of archaea, bacteria and fungi inhabiting intermittent streambed sediment (surface and hyporheic) and buried leaves. Hydrological alterations modified the archaeal community composition more than the bacterial community composition, whereas fungi were the least affected. Throughout the experiment, archaeal communities colonizing sediments showed greater phylogenetic distances compared to those of bacteria and fungi, suggesting considerable adaptation to severe hydrological disturbances. The increase in the class abundances, such as those of Thermoplasmata within archaea and of Actinobacteria and Bacilli within bacteria, revealed signs of transitioning to a drought-favoured and soil-like community composition. Strikingly, we found that in comparison to the drying phase, water return (as sporadic storms and rewetting) led to larger shifts in the surface microbial community composition and diversity. In addition, microhabitat characteristics, such as the greater capacity of the hyporheic zone to maintain/conserve moisture, tended to modulate the ability of certain microbes (e.g., bacteria) to cope with severe hydrological disturbances.