Impact of Protozoan Grazing on Nitrification and the Ammonia-And-Nitrite-Oxidizing Bacterial Communities in Activated Sludge

The effect of protozoan grazing on nitrification rates under different conditions was examined. The spatial distribution of ammonia -and nitrite- oxidizing bacteria (AOB and NOB) in activated sludge was also examined using FISH/CSLM. Batch reactors were monitored for ammonia, nitrite, nitrate, and total nitrogen concentrations and bacterial numbers in the presence and absence of cycloheximide, a protozoan inhibitor. In the absence of protozoan grazing, rates of nitrification were lower than in batches with protozoa. Spatially, both AOB and NOB were found clustered within the floc and neither inhibiting the protozoa or inhibiting ammonia oxidation appeared to lower the amount of AOB and NOB present or their position. These results suggest that a reduction in protozoan grazing pressure allowed the heterotrophic bacteria to proliferate which caused a corresponding decrease in the rate of nitrification. These results suggest that AOB and NOB are less active in the absence of protozoa and indicates the role of protozoa in the cycling of nitrogen.

Impact of Protozoan Grazing on Nitrification and the Ammonia-And-Nitrite-Oxidizing Bacterial Communities in Activated Sludge

The effect of protozoan grazing on nitrification rates under different conditions was examined. The spatial distribution of ammonia -and nitrite- oxidizing bacteria (AOB and NOB) in activated sludge was also examined using FISH/CSLM. Batch reactors were monitored for ammonia, nitrite, nitrate, and total nitrogen concentrations and bacterial numbers in the presence and absence of cycloheximide, a protozoan inhibitor. In the absence of protozoan grazing, rates of nitrification were lower than in batches with protozoa. Spatially, both AOB and NOB were found clustered within the floc and neither inhibiting the protozoa or inhibiting ammonia oxidation appeared to lower the amount of AOB and NOB present or their position. These results suggest that a reduction in protozoan grazing pressure allowed the heterotrophic bacteria to proliferate which caused a corresponding decrease in the rate of nitrification. These results suggest that AOB and NOB are less active in the absence of protozoa and indicates the role of protozoa in the cycling of nitrogen.

Pseudomonas chlororaphis PA23 metabolites protect against protozoan grazing by the predator Acanthamoeba castellanii

Background Pseudomonas chlororaphis strain PA23 is a biocontrol agent that is able to protect canola against the pathogenic fungus Sclerotinia sclerotiorum. This bacterium secretes a number of metabolites that contribute to fungal antagonism, including pyrrolnitrin (PRN), phenazine (PHZ), hydrogen cyanide (HCN) and degradative enzymes. In order to be successful, a biocontrol agent must be able to persist in the environment and avoid the threat of grazing predators. The focus of the current study was to investigate whether PA23 is able to resist grazing by the protozoan predator Acanthamoeba castellanii (Ac) and to define the role of bacterial metabolites in the PA23-Ac interaction. Methods Ac was co-cultured with PA23 WT and a panel of derivative strains for a period of 15 days, and bacteria and amoebae were enumerated on days 1, 5, 10 and 15. Ac was subsequently incubated in the presence of purified PRN, PHZ, and KCN and viability was assessed at 24, 48 and 72 h. Chemotactic assays were conducted to assess whether PA23 compounds exhibit repellent or attractant properties towards Ac. Finally, PA23 grown in the presence and absence of amoebae was subject to phenotypic characterization and gene expression analyses. Results PRN, PHZ and HCN were found to contribute to PA23 toxicity towards Ac trophozoites, either by killing or inducing cyst formation. This is the first report of PHZ-mediated toxicity towards amoebae. In chemotaxis assays, amoebae preferentially migrated towards regulatory mutants devoid of extracellular metabolite production as well as a PRN mutant, indicating this antibiotic has repellent properties. Co-culturing of bacteria with amoebae led to elevated expression of the PA23 phzI/phzR quorum-sensing (QS) genes and phzA and prnA, which are under QS control. PHZ and PRN levels were similarly increased in Ac co-cultures, suggesting that PA23 can respond to predator cues and upregulate expression of toxins accordingly. Conclusions PA23 compounds including PRN, PHZ and HCN exhibited both toxic and repellent effects on Ac. Co-culturing of bacteria and amoebae lead to changes in bacterial gene expression and secondary metabolite production, suggesting that PA23 can sense the presence of these would-be predators and adjust its physiology in response.

Elevated Contribution of Low Nucleic Acid Prokaryotes and Viral Lysis to the Prokaryotic Community Along the Nutrient Gradient From an Estuary to Open Ocean Transect

Prokaryotes represent the largest living biomass reservoir in aquatic environments and play a crucial role in the global ocean. However, the factors that shape the abundance and potential growth rate of the ecologically distinct prokaryotic subgroups [i.e., high nucleic acid (HNA) and low nucleic acid (LNA) cells] along varying trophic conditions in the ocean remain poorly understood. This study conducted a series of modified dilution experiments to investigate how the abundance and potential growth rate of HNA and LNA prokaryotes and their regulating factors (i.e., protozoan grazing and viral lysis) change along a cross-shore nutrient gradient in the northern South China Sea. The results showed that the abundance of both HNA and LNA cells was significantly positively correlated with the abundance of heterotrophic nanoflagellates and viruses, whereas only HNA abundance exhibited a significant positive correlation with nutrient level. With a decreasing nutrient concentration, the potential growth rate of the HNA subgroup declined significantly, while that of the LNA subgroup was significantly enhanced, leading to an elevated relative potential growth rate of the LNA to HNA subgroup under decreasing nutrient levels. Furthermore, our data revealed different regulatory roles of protozoan grazing and viral lysis on the HNA and LNA subgroups, with HNA suffering higher mortality pressure from grazing than from lysis in contrast to LNA, which experienced equivalent pressures. As the nutrient levels declined, the relative contribution of lysis to the mortality of the HNA subgroup increased significantly, in contrast to the insignificant change in that of the LNA subgroup. Our results indicated the elevated role of LNA cells in the prokaryotic community and the enhanced viral lysis pressure on the total prokaryotes under oligotrophic conditions. This implies a weakened efficiency of carbon cycling within the microbial loop and enhanced viral lysis to shunt more carbon and energy flow in the future ocean, in which oligotrophication will be strengthened due to global warming.

Intracellular Metabolites in Marine Microorganisms during an Experiment Evaluating Microbial Mortality

Metabolomics is a tool with immense potential for providing insight into the impact of biological processes on the environment. Here, we used metabolomics methods to characterize intracellular metabolites within marine microorganisms during a manipulation experiment that was designed to test the impact of two sources of microbial mortality, protozoan grazing and viral lysis. Intracellular metabolites were analyzed with targeted and untargeted mass spectrometry methods. The treatment with reduced viral mortality showed the largest changes in metabolite concentrations, although there were organic compounds that shifted when the impact of protozoan grazers was reduced. Intracellular concentrations of guanine, phenylalanine, glutamic acid, and ectoine presented significant responses to changes in the source of mortality. Unexpectedly, variability in metabolite concentrations were not accompanied by increases in microbial abundance which indicates that marine microorganisms altered their internal organic carbon stores without changes in biomass or microbial growth. We used Weighted Correlation Network Analysis (WGCNA) to identify correlations between the targeted and untargeted mass spectrometry data. This analysis revealed multiple unknown organic compounds were correlated with compatible solutes, also called osmolytes or chemical chaperones, which emphasizes the dominant role of compatible solutes in marine microorganisms.

A community-based approach to identifying defence of microalgae against protozoan grazing

It has increasingly been recognized that defence of microalgae against predator grazing is a passive response to increase algal population density by excreting chemicals with a change in physical properties. As common biological pollutants in the cultivation of the microalgae, the community-based method was used to identify the ability of two microalgae, Chlorella sp. and Nannochloropsis oceanica, to defend against protozoan grazing. Mature protozoan samples with 14-day age were collected, using microscopy glass slides, in coastal waters of the Yellow Sea, northern China. For both microalgae, a gradient of concentrations was designed as 100 (control), 104, 105, 106 and 107 cell ml−1, respectively. Results showed that both test algal species represented strong defence effects on protozoan grazing, especially at high density levels. Species richness, abundance and taxonomic distinctness of the protozoan assemblages showed a sharp decrease at high concentration level (107 cell ml−1) of both algae. A significant variation in protozoan community structures was found to be driven by the gradient of the algal concentrations. The paired taxonomic distinctness indices of the protozoan communities showed an increasing trend of departure from the expected taxonomic pattern with increase of algal concentrations. Based on the results, we suggest that the community-based bioassay might be used as a feasible tool for identifying defence against protozoan grazing of microalgae.