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Aerobic granular sludge is a compact and efficient biofilm process used for wastewater treatment which has received much attention and is currently being implemented worldwide. The microbial associations and their ecological implications occurring during granule development, especially those involving inter-kingdom interactions, are poorly understood. In this work, we monitored the prokaryote and eukaryote community composition and structure during the granulation of activated sludge for 343 days in a sequencing batch reactor (SBR) and investigated the influence of abiotic and biotic factors on the granule development. Sludge granulation was accomplished with low-wash-out dynamics at long settling times, allowing for the microbial communities to adapt to the SBR environmental conditions. The sludge granulation and associated changes in microbial community structure could be divided into three stages: floccular, intermediate, and granular. The eukaryotic and prokaryotic communities showed parallel successional dynamics, with three main sub-communities identified for each kingdom, dominating in each stage of sludge granulation. Although inter-kingdom interactions were shown to affect community succession during the whole experiment, during granule development random factors like the availability of settlement sites or drift acquired increasing importance. The prokaryotic community was more affected by deterministic factors, including reactor conditions, while the eukaryotic community was to a larger extent shaped by biotic interactions (including inter-kingdom interactions) and stochasticity., (© 2024. The Author(s).)

Granular sludge is an efficient and compact biofilm process for wastewater treatment. However, the ecological factors involved in microbial community assembly during the granular biofilm formation are poorly understood, and little is known about the reproducibility of the process. Here, three replicate bioreactors were used to investigate microbial succession during the formation of granular biofilms. We identified three successional phases. During the initial phase, the successional turnover was high and α-diversity decreased as a result of the selection of taxa adapted to grow on acetate and form aggregates. Despite these dynamic changes, the microbial communities in the replicate reactors were similar. The second successional phase occurred when the settling time was rapidly decreased to selectively retain granules in the reactors. The influence of stochasticity on succession increased and new niches were created as granules emerged, resulting in temporarily increased α-diversity. The third successional phase occurred when the settling time was kept stable and granules dominated the biomass. Turnover was low, and selection resulted in the same abundant taxa in the reactors, but drift, which mostly affected low-abundant community members, caused the community in one reactor to diverge from the other two. Even so, performance was stable and similar between reactors.

Granular activated sludge has gained increasing interest due to its potential in treating wastewater in a compact and efficient way. It is well-established that activated sludge can form granules under certain environmental conditions such as batch-wise operation with feast-famine feeding, high hydrodynamic shear forces, and short settling time which select for dense microbial aggregates. Aerobic granules with stable structure and functionality have been obtained with a range of different wastewaters seeded with different sources of sludge at different operational conditions, but the microbial communities developed differed substantially. In spite of this, granule instability occurs. In this review, the available literature on the mechanisms involved in granulation and how it affects the effluent quality is assessed with special attention given to the microbial interactions involved. To be able to optimize the process further, more knowledge is needed regarding the influence of microbial communities and their metabolism on granule stability and functionality. Studies performed at conditions similar to full-scale such as fluctuation in organic loading rate, hydrodynamic conditions, temperature, incoming particles, and feed water microorganisms need further investigations.

Microorganisms colonize surfaces and develop biofilms through interactions that are not yet thoroughly understood, with important implications for water and wastewater systems. This study investigated the interactions between N-acyl homoserine lactone (AHL)-producing bacteria, yeasts and protists, and their contribution to biofilm development. Sixty-one bacterial strains were isolated from activated sludge and screened for AHL production, with Aeromonas sp. found to be the dominant AHL producer. Shewanella xiamenensis, Aeromonas allosaccharophila, Acinetobacter junii and Pseudomonas aeruginosa recorded the highest adherence capabilities, with S. xiamenensis being the most effective in surface colonization. Additionally, highly significant interactions (i.e. synergic or antagonistic) were described for dual and multistrain mixtures of bacterial strains (P. aeruginosa, S. xiamenensis, A. junii and Pseudomonas stutzeri), as well as for strongly adherent bacteria co-cultured with yeasts. In this last case, the adhered biomass in co-cultures was lower than the monospecific biofilms of bacteria and yeast, with biofilm observations by microscopy suggesting that bacteria had an antagonist effect on the whole or part of the yeast population. Finally, protist predation by Euplotes sp. and Paramecium sp. on Aeromonas hydrophila biofilms not only failed to reduce biofilm formation, but also recorded unexpected results leading to the development of aggregates of high density and complexity., (© FEMS 2016. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

The quality of the sludge in Wastewater Treatment Plants (WWTPs) depends on the suitable colonization of the flocs by microorganisms. Due to the functional importance of these biological constituents, several biological or biological-related parameters have been commonly used for the control of depuration efficiency. According to national and international water regulation recommendations, interlaboratory studies have a great relevance to determine which parameters are more reliable for their extensive application in routine control. However, these studies are also very useful to demonstrate consistency in results from multiple laboratories and to develop reliable and reproducible methodologies which might be necessary for protocol validation and also for accreditation issues to meet regulatory environmental requirements. The main purpose of this work was to assess the results obtained in consecutive interlaboratory assays in order to determine the concordance degree in the application of biological parameters by participating laboratories. Following the international recommendations about these studies, a common working protocol was proposed. Statistical tests indicated that Sludge Index and several routine physical-chemical analyses [V30, Mixed Liquor Suspended Solids (MLSS), Mixed Liquor Volatile Solids (MLVS) and Sludge Volumetric Index (SVI)] show low variability and therefore are suitable tools for laboratory control. Shannon Index and Sludge Biotic Index also presented low variability although a more precise protocol would be necessary, in particular the methodology to count small flagellates. The abundance and identification of protist species showed low concordance among laboratories and three factors were responsible for the low reliability of data: population density, size and morphological distinguishable characters of the specimens.