Evaluating CO2 emissions from continuous flow and batch growth systems under autotrophic mode: Implications for GHG accounting of biological nutrient removal.

The oxidation of ammonia by autotrophic bacteria is a central part of the nitrogen cycle and a fundamental aspect of biological nutrient removal (BNR) during wastewater treatment. Autotrophic ammonia oxidation produces protons and results in net-CO2 production due to the neutralizing effect of bicarbonate alkalinity. Attention must be paid to the propensity for this produced CO2 to be transferred to the atmosphere where it can act as a greenhouse gas (GHG). In the context of BNR systems, bicarbonate-derived CO2 emissions should be considered distinct from the biogenic CO2 that arises from cellular respiration, though this distinction is not made in current GHG accounting practices. The aim of this study was to evaluate the performance of two experimental systems operated under autotrophic mode and buffered with bicarbonate, to investigate the relationship between ammonia removal and gaseous CO2 emissions. The first system consisted of continuously aerated lab-scale batch reactors, which were effective in demonstrating the important link between ammonia oxidizer activity, pH, and gaseous CO2 production. Depletion of the buffer system always led to a rapid decline in system pH and cessation of CO2 emissions when the pH fell below 7.0. The second system was a tubular continuous-flow biofilm reactor which permitted comparison of ammonia removal and CO2 emission rates. A linear relationship between ammonia removal and CO2 emissions was demonstrated and the quantified CO2 production was relatively close to that which was predicted based on the stoichiometry of nitrification, with this CO2 being detected in the gas phase. It was apparent that this system offered minimal resistance to the mass transfer of CO2 from the liquid to gas, which is an important factor that determines how much of the bicarbonate-derived CO2 may contribute to greenhouse gas emissions in engineered systems such as those used for BNR.

Recent advancements in the biological treatment of high strength ammonia wastewater.

The estimated global population growth of 81 million people per year, combined with increased rates of urbanization and associated industrial processes, result in volumes of high strength ammonia wastewater that cannot be treated in a cost-effective or sustainable manner using the floc-based conventional activated sludge approach of nitrification and denitrification. Biofilm and aerobic granular sludge technologies have shown promise to significantly improve the performance of biological nitrogen removal systems treating high strength wastewater. This is partly due to enhanced biomass retention and their ability to sustain diverse microbial populations with juxtaposing growth requirements. Recent research has also demonstrated the value of hybrid systems with heterogeneous bioaggregates to mitigate biofilm and granule instability during long-term operation. In the context of high strength ammonia wastewater treatment, conventional nitrification-denitrification is hampered by high energy costs and greenhouse gas emissions. Anammox-based processes such as partial nitritation-anammox and partial denitrification-anammox represent more cost-effective and sustainable methods of removing reactive nitrogen from wastewater. There is also growing interest in the use of photosynthetic bacteria for ammonia recovery from high strength waste streams, such that nitrogen can be captured and concentrated in its reactive form and recycled into high value products. The purpose of this review is to explore recent advancements and emerging approaches related to high strength ammonia wastewater treatment.

Interspecies interaction extends bacterial survival at solid-air interfaces.

Despite the ubiquity of biofilms in natural and man-made environments, research on surface-associated cells has focused primarily on solid-liquid interfaces. This study evaluated the extent to which bacterial cells persist on inanimate solid-air interfaces. The desiccation tolerance of bacterial strains isolated from indoor air, as well as of a test strain (Pseudomonas aeruginosa), was determined at different levels of relative humidity (RH) using the large droplet inoculation method in an aerosol chamber. The cells survived longer at lower (25 and 42%) than at high RH (95%). Four of the seven indoor strains selected for further study showed extended period of survival following deposition as 0.05-0.1 ml of washed culture followed by desiccation, each with different effects on the survival of the test strain, P. aeruginosa. A strain closely related to Arthrobacter species afforded the highest level of protection to the test strain. Even though the desiccation-tolerant strains survived when they were deposited as bioaerosols, the protective role towards the test strain was not observed when the latter was deposited as a bioaerosol. These, which are often-unculturable, bacteria may go undetected during routine monitoring of biofouling, thereby allowing them to act as reservoirs and extending the habitat range of undesired microorganisms.

Huddling together to survive: Population density as a survival strategy of non-spore forming bacteria under nutrient starvation and desiccation at solid-air interfaces.

Acclimation and flexible response mechanisms are survival adaptations allowing prokaryotic cells to colonize diverse habitats and maintain viability in nature. Lack of water significantly impacts cellular response, which can be partially compensated for through community interactions and accessing survival means beyond the cell's boundaries. In the present study, higher numbers of cultivable Gram-positive Arthrobacter sp. and Gram-negative Pseudomonas stutzeri cells were found on surfaces when high population density was used after prolonged periods of desiccation and nutrient starvation. Total cell counts during desiccation periods decreased slower than culturable cell counts independently from initial population density. The presence of homogenate, prepared by filtering homogenized cultures through a 0.2 µm filter, extended culturability of Arthrobacter sp. cells, while intact heat-killed cells extended the culturability of Arthrobacter sp. and P. stutzeri. Our results suggest very slow cell membrane breakdown for desiccated bacterial cells at solid-air interfaces over extended time spans, which may serve as reservoirs of nutrients, and may potentially provide trace amounts of water for surviving cells. Higher initial population density and recycling of resources from "zombie"-like cells, may support growth in a similar fashion as access to cell lysates or the contents of heat-killed cells analogous to dead-phase cultures where some cells experience cryptic growth.

Species Interaction and Selective Carbon Addition During Antibiotic Exposure Enhances Bacterial Survival.

Biofilms are multifaceted and robust microbiological systems that enable microorganisms to withstand a multitude of environmental stresses and expand their habitat range. We have shown previously that nutritional status alters antibiotic susceptibility in a mixed-species biofilm. To further elucidate the effects of nutrient addition on inter-species dynamics and whole-biofilm susceptibility to high-dose streptomycin exposures, a CO2 Evolution Measurement System was used to monitor the metabolic activity of early steady state pure-culture and mixed-species biofilms containing Pseudomonas aeruginosa and Stenotrophomonas maltophilia, with and without added carbon. Carbon supplementation was needed for biofilm recovery from high-dose streptomycin exposures when P. aeruginosa was either the dominant community member in a mixed-species biofilm (containing predominantly P. aeruginosa and S. maltophilia) or as a pure culture. By contrast, S. maltophilia biofilms could recover from high-dose streptomycin exposures without the need for carbon addition during antibiotic exposure. Metagenomic analysis revealed that even when inocula were dominated by Pseudomonas, the relative abundance of Stenotrophomonas increased upon biofilm development to ultimately become the dominant species post-streptomycin exposure. The combined metabolic and metagenomic results demonstrated the relevance of inter-species influence on survival and that nutritional status has a strong influence on the survival of P. aeruginosa dominated biofilms.

Surface-attached sulfonamide containing quaternary ammonium antimicrobials for textiles and plastics.

With the risks associated with healthcare-associated infections and the rise of antibiotic resistant microorganisms, there is an important need to control the proliferation of these factors in hospitals, retirement homes and other institutions. This work explores the development and application of a novel class of sulfonamide-based quaternary ammonium antimicrobial coatings, anchored to commercially and clinically relevant material surfaces. Synthesized in high yields (60-97%), benzophenone-anchored antimicrobials were spray-coated and UV grafted onto plastic surfaces, while silane-anchored variants were adhered to select textiles via dip-coating. Surface modified samples were characterised by advancing contact angle, anionic dye staining, X-ray photoelectron spectroscopy and atomic force microscopy. After verifying coating quality through the above characterization methods, microbiological testing was performed on batch samples in conditions that simulate the natural inoculation of surfaces and objects (solid/air) and water containers (solid/liquid). Using the previously established Large Drop Inoculum (LDI) protocol at solid/air interfaces, all treated samples showed a full reduction (105-107 CFU) of viable Arthrobacter sp., S. aureus, and E. coli after 3 h of contact time. Additional testing of the walls of plastic LDPE vials treated with a UV-cured sulfonamide antimicrobial at a solid/liquid interface using the newly developed Large Reservoir Inoculum (LRI) protocol under static conditions revealed a complete kill (>106 reduction) of Gram-positive Arthrobacter sp., and a partial kill (>104 reduction) of Gram-negative E. coli within 24-48 h of contact.

UV-Curable Contact Active Benzophenone Terminated Quaternary Ammonium Antimicrobials for Applications in Polymer Plastics and Related Devices.

A series of UV active benzophenone ([C6H5COC6H4-O-(CH2)n-N+Me2R][X-]; 4, R = C12H25, n = 3, X- = Br-; 5a-c, R = C18H37, n = 3, X- = Cl-, Br-, I-; 6a-c, R = C18H37, n = 4, X- = Cl-, Br-, I-; 7a-c, R = C18H37, n = 6, X- = Cl-, Br-, I-) terminated C12 and C18 quaternary ammonium salts (QACs) were prepared by thermal or microwave-driven Menshutkin protocols of the appropriate benzophenone alkyl halide (1a-c, 2a-c, 3a-c) with the corresponding dodecyl- or octadecyl N,N-dimethylamine. All new compounds were characterized by NMR spectroscopy, HRMS spectrometry, and, in one instance (4), by single-crystal X-ray crystallography. Representative C12 and C18 benzophenone QACs were formulated into 1% (w/v) water or water/ethanol-based aerosol spray coatings and then UV-cured onto plastic substrates (polypropylene, polyethylene, polystyrene, polyvinyl chloride, and polyether ether ketone) with exposure to low to moderate doses of UV (20-30 J cm-2). Confirmation as to the presence of the coatings was detected by advancing water contact angle measurements, which revealed a more hydrophilic surface after coating. Further confirmation was gained by X-ray photoelectron spectroscopy analysis, time of flight secondary ion mass spectrometry, and bromophenol blue staining, all of which showed the presence of the attached quaternary ammonium molecule. Analysis of surfaces treated with the C18 benzophenone 5b by atomic force microscopy and surface profilometry revealed a coating thickness of ∼350 nm. The treated samples along with controls were then evaluated for their antimicrobial efficacy against Gram-positive (Arthrobacter sp., Listeria monocytogenes) and Gram-negative (Pseudomonas aeruginosa) bacteria at a solid/air interface using the large drop inoculum protocol; this technique gave no evidence for cell adhesion after a 3 h time frame. These antimicrobial materials show promise for their use as coatings on plastic biomedical devices with the aim of preventing biofilm formation and preventing the spread of hospital acquired infections.

USMB-induced synergistic enhancement of aminoglycoside antibiotics in biofilms.

This study evaluated the effect of combining antibiotics with ultrasound and microbubbles (USMB) toward the eradication of biofilms. Pseudomonas aeruginosa PAO1 biofilms were treated with the antibiotics gentamicin sulfate or streptomycin sulfate, or a combination of USMB with the respective antibiotics. Biofilm structure was quantified using confocal laser scanning microscopy with COMSTAT analysis, while activity was measured as whole-biofilm CO2 production in a continuous-flow biofilm model. The combined antibiotic-USMB treatment significantly impacted biofilm biomass, thickness and surface roughness compared to antibiotics alone (p<0.05). USMB exposure caused the formation of craters (5-20µm in diameter) in the biofilms, and when combined with gentamicin, activity was significantly lower, compared to gentamicin, USMB or untreated controls, respectively. Interestingly, the CO2 production rate following combined streptomycin-USMB treatment was higher than after streptomycin alone, but significantly lower than USMB alone and untreated control. These results show strong evidence of a synergistic effect between antibiotics and USMB, although the varied response to different antibiotics emphasize the need to optimize the USMB exposure conditions to maximize this synergism and ultimately transfer this technology into clinical or industrial practice.