Abstract
River bacterioplankton communities, influenced by watershed usage, are responsible for water purification. Bacterioplankton may be critical in the degradation of dissolved organic nitrogen: the major nitrogen pool in the Caloosahatchee River, Florida. We investigated how freshwater discharge influences estuarine bacterioplankton and how the freshwater-originated DON is utilized by estuarine bacterioplankton. Microcosm experiments were conducted during low and high discharge using two upstream freshwater samples: one site primarily influenced by Lake Okeechobee and the other site moderately influenced by an agricultural watershed. These freshwater samples were filtered to eliminate indigenous microbial populations, then mixed with estuarine bacterioplankton. High-throughput sequencing revealed that bacterioplankton differed between low and high discharge and were influenced by salinity. Alphaproteobacteria and Bacteroidetes dominated in low discharge while Bacteroidetes and Cyanobacteria dominated during high discharge. In the microcosm experiment, DON concentration decreased with increasing cell densities, suggesting that the DON was utilized as a carbon and nitrogen source. Band signals in denaturing gradient gel electrophoresis corresponding to Alphaproteobacteria and Actinobacteria decreased while Gammaproteobacteria increased during the 1 month incubation. This data suggests that estuarine bacterioplankton communities are influenced by variations in discharge patterns and use freshwater-originated DON as demonstrated by a shift in community structure. Bacteria in the genus Nitrosospira play the vital role of converting ammonia to nitrite via the enzymes ammonia monooxygenase and hydroxylamine oxidoreductase. Nitrosospira lacus strain APG3, isolated from Green Lake, Seattle, WA is a psychrotolerant betaproteobacterial ammonia-oxidizing bacterium that can grow at temperatures as low as 4°C. Nitrosospira lacus is able to grow over a wide pH range (5-9) and like other Nitrosospira, is susceptible to high ammonium concentrations. APG3 showed the ability to grow at low pH and use urea which may allow it to survive more efficiently in soil environments. 16S rRNA analysis revealed that the closest relative of APG3 is Nitrosospira multiformis (cluster 3). When using ANI values to compare APG3 to Nitrosospira multiformis, the ANI value demonstrates that they are related but different species that share less than half of their genome. Nitrosospira lacus is the first isolated cluster 0 representative. The distribution of Nitrosospira lacus like sequences reveals a habitat limitation to tropical and subtropical areas with a preference for waterlogged areas. The draft genome sequence comprises 3,107,181 bases at 272-fold coverage. The assembled draft genome consists of 84 contigs with an average size of 41,181 bp and a G+C content of 53.6%. The genome contains 3,147 protein-coding DNA sequences, 44 tRNA genes, and a single 16S-23S-5S rRNA operon. The genome revealed that Nitrosospira lacus represents a new species of cluster 0 Nitrosospira, which contains a slew of genes that allow it to incorporate nutrients from the environment. The genome of N. lacus when compared to other ammonia-oxidizing bacteria most closely resembles that of N. multiformis.The presence of hydrogenase and urease, suggests that Nitrosospira lacus may be able to meet its energy needs via alternative pathways. N. lacus also contains genes to cope with various degrees of stress, including detoxification, cold and heat shock proteins and oxidative and osmotic stress, copper, cobalt, zinc, and cadmium resistance. This unique genetic makeup may allow Nitrosospira lacus adaptability to a wide range of freshwater habitats.