Microbial community-based protein from soybean-processing wastewater as a sustainable alternative fish feed ingredient.

As the global demand for food increases, aquaculture plays a key role as the fastest growing animal protein sector. However, existing aquafeeds contain protein ingredients that are not sustainable under current production systems. We evaluated the use of microbial community-based single cell protein (SCP), produced from soybean processing wastewater, as a partial fishmeal protein substitute in juvenile Asian seabass (Lates calcarifer). A 24-day feeding trial was conducted with a control fishmeal diet and a 50% fishmeal replacement with microbial community-based SCP as an experimental group, in triplicate tanks containing 20 fish each. Both diets met the protein, essential amino acids (except for lysine), and fat requirements for juvenile Asian sea bass. The microbial composition of the SCP was dominated by the genera Acidipropionibacterium and Propioniciclava, which have potential as probiotics and producers of valuable metabolites. The growth performance in terms of percent weight gain, feed conversion ratio (FCR), specific growth rate (SGR), and survival were not significantly different between groups after 24 days. The experimental group had less variability in terms of weight gain and FCR than the control group. Overall, microbial community-based protein produced from soybean processing wastewater has potential as a value-added feed ingredient for sustainable aquaculture feeds.

Metagenomics and metatranscriptomics suggest pathways of 3-chloroaniline degradation in wastewater reactors.

Biological wastewater treatment systems are often affected by shifts in influent quality, including the input of toxic chemicals. Yet the mechanisms underlying the adaptation of activated sludge process performance are rarely studied in a controlled and replicated experimental setting, particularly when challenged with a sustained toxin input. Three replicate bench-scale bioreactors were subjected to a chemical disturbance in the form of 3-chloroaniline (3-CA) over 132 days, after an acclimation period of 58 days, while three control reactors received no 3-CA input. Ammonia oxidation was initially affected by 3-CA. Within three weeks of the experiment, microbial communities in all three treatment reactors adapted to biologically degrade 3-CA resulting in partial ammonia oxidation recovery. Combining process and microbial community data from amplicon sequencing with potential functions gleaned from assembled metagenomics and metatranscriptomics data, two putative degradation pathways for 3-CA were identified. The first pathway, determined from metagenomics data, involves a benzoate dioxygenase and subsequent meta-cleavage of the aromatic ring. The second, determined from intensive short-term sampling for gene expression data in tandem with 3-CA degradation, involves a phenol monooxygenase followed by ortho-cleavage of the aromatic ring. The relative abundances of amplicon sequence variants associated with the genera Gemmatimonas, OLB8, and Taibaiella correlated significantly with 3-CA degradation. Metagenome-assembled genome data also showed the genus OLB8 to be differentially enriched in treatment reactors, making it a strong candidate as 3-CA degrader. Using replicated reactors, this study has demonstrated the impact of a sustained stress on the activated sludge process. The unique and novel features of this study include the identification of putative pathways and potential degraders of 3-CA using long-term and short-term sampling in tandem with multiple methods in a controlled and replicated experiment.

Microbial community-based production of single cell protein from soybean-processing wastewater of variable chemical composition.

The use of food-processing wastewaters to produce microbial biomass-derived single cell protein (SCP) is a sustainable way to meet the global food demand. Microbial community-based approaches to SCP production have the potential benefits of lower costs and greater resource recovery compared to pure cultures, yet they have received scarce attention. Here, SCP production from soybean-processing wastewaters using their existent microbial communities was evaluated. Six sequencing batch reactors of 4.5-L working volume were operated at 30 °C for 34 d in cycles consisting of 3-h anaerobic and 9-h aerobic phases. Four reactors received no microbial inoculum and the remaining two were amended with 1.5 L of a mixed culture from a prior SCP production cycle. Reactors produced more SCP when fed with wastewaters of higher soluble total Kjeldahl nitrogen (sTKN) content. The protein yield in biomass ranged from 0.53 to 3.13 g protein/g sTKN, with a maximum protein content of 50 %. The average removal of soluble chemical oxygen demand (sCOD) and soluble total nitrogen (sTN) was 92 % and 73 %, respectively. Distinct microbial genera were enriched in all six bioreactors, with Azospirillum, Rhodobacter, Lactococcus, and Novosphingobium dominating. The study showed that constituents in soybean wastewater can be converted to SCP and demonstrated the effect of variable influent wastewater composition on SCP production.

Microbiome assembly predictably shapes diversity across a range of disturbance frequencies in experimental microcosms.

Diversity is often implied to have a positive effect on the functional stability of ecological communities. However, its relationship with stochastic and deterministic assembly mechanisms remains largely unknown, particularly under fluctuating disturbances. Here, we subjected complex bacterial communities in microcosms to different frequencies of alteration in substrate feeding scheme, tracking temporal dynamics in their assembly, structure and function. Activated sludge bioreactors were subjected to six different frequencies of double organic loading, either never (undisturbed), every 8, 6, 4, or 2 days (intermediately disturbed), or every day (press disturbed), and operated in daily cycles for 42 days. Null modeling revealed a stronger role of stochastic assembly at intermediate disturbance frequencies, with a peak in stochasticity that preceded the occurrence of a peak in α-diversity. Communities at extreme ends of the disturbance range had the lowest α-diversity and highest within-treatment similarity in terms of ß-diversity, with stronger deterministic assembly. Increased carbon removal and microbial aggregate settleability (general functions) correlated with stronger deterministic processes. In contrast, higher stochasticity correlated with higher nitrogen removal (a specialized function) only during initial successional stages at intermediate disturbance frequencies. We show that changes in assembly processes predictably precede changes in diversity under a gradient of disturbance frequencies, advancing our understanding of the mechanisms behind disturbance-diversity-function relationships.

Microbial community-based protein production from wastewater for animal feed applications.

Single cell protein (SCP) derived from microbial biomass represents a promising source of protein for animal feed additives. While microbial community-based approaches to SCP production using nutrient-rich wastewaters incur lower costs than traditional single organism-based approaches, they have received little attention. This review focuses on SCP production using wastewaters with an emphasis on food-processing wastewaters. An elemental carbon-to-nitrogen ratio ranging from 10 to 20 is recommended to promote a high microbial biomass protein yield. Proteobacteria was identified as the most prevalent phylum within SCP-producing microbial communities. More research is needed to determine the composition of the microbial community best suited for SCP production, as well as its relationship with the microbial community in influent food-processing wastewaters. Remaining challenges are target protein and essential amino acids content, protein quantification and biomass yield assessment. The review presents bioreactor design considerations towards defining suitable operating conditions for SCP production through microbial community-based fermentation.

Press Disturbance Alters Community Structure and Assembly Mechanisms of Bacterial Taxa and Functional Genes in Mesocosm-Scale Bioreactors.

Press disturbances are of interest in microbial ecology, as they can drive microbial communities to alternative stable states. However, the effect of press disturbances in community assembly mechanisms, particularly with regard to taxa and functional genes at different levels of abundance (i.e., common and rare), remains largely unknown. Here, we tested the effect of a continuous alteration in substrate feeding scheme on the structure, function, and assembly of bacterial communities. Two sets of replicate 5-liter sequencing batch reactors were operated at two different organic carbon loads for a period of 74 days, following 53 days of acclimation after inoculation with sludge from a full-scale treatment plant. Temporal dynamics of community taxonomic and functional gene structure were derived from metagenomics and 16S rRNA gene metabarcoding data. Disturbed reactors exhibited different community function, structure, and assembly compared to undisturbed reactors. Bacterial taxa and functional genes showed dissimilar α-diversity and community assembly patterns. Deterministic assembly mechanisms were generally stronger in disturbed reactors and in common fractions compared to rare ones. Function quickly recovered after the disturbance was removed, but community structure did not. Our results highlight that functional gene data from metagenomics can indicate patterns of community assembly that differ from those obtained from taxon data. This study reveals how a joint evaluation of assembly mechanisms and community structure of bacterial taxa and functional genes as well as ecosystem function can unravel the response of complex microbial systems to a press disturbance.IMPORTANCE Ecosystem management must be viewed in the context of increasing frequencies and magnitudes of various disturbances that occur at different scales. This work provides a glimpse of the changes in assembly mechanisms found in microbial communities exposed to sustained changes in their environment. These mechanisms, deterministic or stochastic, can cause communities to reach a similar or variable composition and function. For a comprehensive view, we use a joint evaluation of temporal dynamics in assembly mechanisms and community structure for both bacterial taxa and their functional genes at different abundance levels, in both disturbed and undisturbed states. We further reverted the disturbance state to contrast recovery of function with community structure. Our findings are relevant, as very few studies have employed such an approach, while there is a need to assess the relative importance of assembly mechanisms for microbial communities across different spatial and temporal scales, environmental gradients, and types of disturbance.

Trait-based life-history strategies explain succession scenario for complex bacterial communities under varying disturbance.

Trait-based approaches are increasingly gaining importance in community ecology, as a way of finding general rules for the mechanisms driving changes in community structure and function under the influence of perturbations. Frameworks for life-history strategies have been successfully applied to describe changes in plant and animal communities upon disturbance. To evaluate their applicability to complex bacterial communities, we operated replicated wastewater treatment bioreactors for 35 days and subjected them to eight different disturbance frequencies of a toxic pollutant (3-chloroaniline), starting with a mixed inoculum from a full-scale treatment plant. Relevant ecosystem functions were tracked and microbial communities assessed through metagenomics and 16S rRNA gene sequencing. Combining a series of ordination, statistical and network analysis methods, we associated different life-history strategies with microbial communities across the disturbance range. These strategies were evaluated using tradeoffs in community function and genotypic potential, and changes in bacterial genus composition. We further compared our findings with other ecological studies and adopted a semi-quantitative competitors, stress-tolerants, ruderals (CSR) classification. The framework reduces complex data sets of microbial traits, functions and taxa into ecologically meaningful components to help understand the system response to disturbance and hence represents a promising tool for managing microbial communities.

Frequency of disturbance alters diversity, function, and underlying assembly mechanisms of complex bacterial communities.

Disturbance is known to affect the ecosystem structure, but predicting its outcomes remains elusive. Similarly, community diversity is believed to relate to ecosystem functions, yet the underlying mechanisms are poorly understood. Here, we tested the effect of disturbance on the structure, assembly, and ecosystem function of complex microbial communities within an engineered system. We carried out a microcosm experiment where activated sludge bioreactors operated in daily cycles were subjected to eight different frequency levels of augmentation with a toxic pollutant, from never (undisturbed) to every day (press-disturbed), for 35 days. Microbial communities were assessed by combining distance-based methods, general linear multivariate models, α-diversity indices, and null model analyses on metagenomics and 16S rRNA gene amplicon data. A stronger temporal decrease in α-diversity at the extreme, undisturbed and press-disturbed, ends of the disturbance range led to a hump-backed pattern, with the highest diversity found at intermediate levels of disturbance. Undisturbed and press-disturbed levels displayed the highest community and functional similarity across replicates, suggesting deterministic processes were dominating. The opposite was observed amongst intermediately disturbed levels, indicating stronger stochastic assembly mechanisms. Trade-offs were observed in the ecosystem function between organic carbon removal and both nitrification and biomass productivity, as well as between diversity and these functions. Hence, not every ecosystem function was favoured by higher community diversity. Our results show that the assessment of changes in diversity, along with the underlying stochastic-deterministic assembly processes, is essential to understanding the impact of disturbance in complex microbial communities.