Activated sludge from municipal wastewater treatment processes can be used directly for the production of biodegradable polyesters from the family of polyhydroxyalkanoates (PHAs). However, municipal activated sludge typically cannot accumulate PHAs to very high levels and often low yields of polymer produced on substrate are observed. In the present work, it was found that the presence of calcium promotes selective growth and enrichment of the PHA-storing biomass fraction and significantly improved both PHA contents and yields. Calcium addition resulted in PHA contents of 0.60 ± 0.03 gPHA/gVSS and average PHA yields on substrate of 0.49 ± 0.03 gCOD_PHA/gCOD_HAc compared to 0.35 ± 0.01 gPHA/gVSS and 0.19 ± 0.01 gCOD_PHA/gCOD_HAc without calcium addition. After 48 h, three times more PHA was produced compared to control experiments without calcium addition. Higher PHA content and selective biomass production is proposed to be a consequence of calcium dependent increased levels of passive acetate uptake. Such more efficient substrate uptake could be related to a formation of calcium acetate complexes. Findings lead to bioprocess methods to stimulate a short-term selective growth of PHA-storing microorganisms and this enables improvements to the techno-economic feasibility for municipal waste activated sludge to become a generic resource for industrial scale PHA production.

Application of animal manure to soils results in the introduction of manure-derived bacteria and their antimicrobial resistance genes (ARGs) into soils. ResCap is a novel targeted-metagenomic approach that allows the detection of minority components of the resistome gene pool without the cost-prohibitive coverage depths and can provide a valuable tool to study the spread of antimicrobial resistance (AMR) in the environment. We used high-throughput sequencing and qPCR for 16S rRNA gene fragments as well as ResCap to explore the dynamics of bacteria, and ARGs introduced to soils and adjacent water ditches, both at community and individual scale, over a period of three weeks. The soil bacteriome and resistome showed strong resilience to the input of manure, as manuring did not impact the overall structure of the bacteriome, and its effects on the resistome were transient. Initially, manure application resulted in a substantial increase of ARGs in soils and adjacent waters, while not affecting the overall bacterial community composition. Still, specific families increased after manure application, either through the input of manure (e.g., Dysgonomonadaceae) or through enrichment after manuring (e.g., Pseudomonadaceae). Depending on the type of ARG, manure application resulted mostly in an increase (e.g., aph(6)-Id), but occasionally also in a decrease (e.g., dfrB3) of the absolute abundance of ARG clusters (FPKM/kg or L). This study shows that the structures of the bacteriome and resistome are shaped by different factors, where the bacterial community composition could not explain the changes in ARG diversity or abundances. Also, it highlights the potential of applying targeted metagenomic techniques, such as ResCap, to study the fate of AMR in the environment.