Individual-based modelling of biofilms accounts for the fact that individual organisms of the same species may well be in a different physiological state as a result of environmental gradients, lag times in responding to change, or noise in gene expression, which we have become increasingly aware of with the advent of single-cell microbiology. But progress in developing and using individual-based modelling has been hampered by different groups writing their own code and the lack of an available standard model. We therefore set out to merge most features of previous models and incorporate various improvements in order to provide a common basis for further developments. Four improvements stand out: the biofilm pressure field allows for shrinking or consolidating biofilms; the continuous-in-time extracellular polymeric substances excretion leads to more realistic fluid behaviour of the extracellular matrix, avoiding artefacts; the stochastic chemostat mode allows comparison of spatially uniform and heterogeneous systems; and the separation of growth kinetics from the individual cell allows condition-dependent switching of metabolism. As an illustration of the model's use, we used the latter feature to study how environmentally fluctuating oxygen availability affects the diversity and composition of a community of denitrifying bacteria that induce the denitrification pathway under anoxic or low oxygen conditions. We tested the hypothesis that the existence of these diverse strategies of denitrification can be explained solely by assuming that faster response incurs higher costs. We found that if the ability to switch metabolic pathways quickly incurs no costs the fastest responder is always the best. However, if there is a trade-off where faster switching incurs higher costs, then there is a strategy with optimal response time for any frequency of environmental fluctuations, suggesting that different types of denitrifying strategies win in different environments. In a single environment, biodiversity of denitrifiers is higher in biofilms than chemostats, higher with than without costs and higher at intermediate frequency of change. The highly modular nature of the new computational model made this case study straightforward to implement, and reflects the sort of novel studies that can easily be executed with the new model.

We present an individual-based experimental framework to identify and estimate the main parameters governing bacterial conjugation at the individual cell scale. From this analysis, we have established that transient periods of unregulated plasmid transfer within newly formed transconjugant cells, together with contact mechanics arising from cellular growth and division, are the two main processes determining the emergent inability of the pWW0 TOL plasmid to fully invade spatially structured Pseudomonas putida populations. We have also shown that pWW0 conjugation occurs mainly at advanced stages of the growth cycle and that nongrowing cells, even when exposed to high nutrient concentrations, do not display conjugal activity. These results do not support previous hypotheses relating conjugation decay in the deeper cell layers of bacterial biofilms to nutrient depletion and low physiological activity. We observe, however, that transient periods of elevated plasmid transfer in newly formed transconjugant cells are offset by unfavorable cell-to-cell contact mechanics, which ultimately precludes the pWWO TOL plasmid from fully invading tightly packed multicellular P. putida populations such as microcolonies and biofilms.

Effective Biological Nitrogen Removal Treatment Processes for Domestic Wastewaters with Low C/N Ratios: A Review Discharge of nitrogenous components to water bodies can cause eutrophication, deterioration of water quality, toxicity to aquatic life, and pose a potential hazard to human and animal health. Biological nitrogen removal can remove nitrogenous components via conversion to harmless nitrogen gas with high efficiency and relative low costs. However, the removal of nitrogen from domestic wastewater with a low carbon/nitrogen (C/N) ratio can often be limited in municipal wastewater plants (WWTPs) because organic carbon is a limiting factor for denitrification. The present work reviews innovative bacterial nitrogen removal pathways such as shortcut nitrification/denitrification, simultaneous nitrification/denitrification, and the nitritation Anammox process, which can remove nitrogen with low or zero dosage of organic carbon sources. We conclude that advanced process control and some new biological treatment processes including the modified anaerobic/anoxic/oxic (A(2)/O) process, the step-feed multistage anaerobic/ oxic (A/O) process, and new reactors like the membrane bioreactors (MBRs) and the membrane-aerated biofilm reactors (MABRs) can support the innovative biological nitrogen removal pathways. They can effectively be used for nitrogen removal from low C/N domestic wastewater without external carbon addition. In addition, conventional and alternative carbon sources for enhanced biological nitrogen removal were also reviewed. We conclude that alternative carbon sources such as wine distillery effluent, the leachate of food waste, digested piggery manure, hydrolyzed molasses, biologically hydrolyzed or mechanically disintegrated sludge offer the same or better performance for nitrogen removal at reduced costs. Finally, we suggest that (1) these new processes and technologies are implemented at large scale for nitrogen removal from low C/N domestic wastewater, (2) further method logic are explored to introduce the Anammox pathway into domestic wastewater treatment, and (3) alternative carbon sources are explored and optimized for supporting the denitrification. With these efforts, cost-effective nitrogen removal from low C/N ratio domestic wastewater can be obtained in the near future.

Plasmid invasion in biofilms is often surprisingly limited in spite of the close contact of cells in a biofilm. We hypothesized that this poor plasmid spread into deeper biofilm layers is caused by a dependence of conjugation on the growth rate (relative to the maximum growth rate) of the donor. By extending an individual-based model of microbial growth and interactions to include the dynamics of plasmid carriage and transfer by individual cells, we were able to conduct in silico tests of this and other hypotheses on the dynamics of conjugal plasmid transfer in biofilms. For a generic model plasmid, we find that invasion of a resident biofilm is indeed limited when plasmid transfer depends on growth, but not so in the absence of growth dependence. Using sensitivity analysis we also find that parameters related to timing (i.e. a lag before the transconjugant can transfer, transfer proficiency and scan speed) and spatial reach (EPS yield, conjugal pilus length) are more important for successful plasmid invasion than the recipients' growth rate or the probability of segregational loss. While this study identifies one factor that can limit plasmid invasion in biofilms, the new individual-based framework introduced in this work is a powerful tool that enables one to test additional hypotheses on the spread and role of plasmids in microbial biofilms.