Adaptation and evolution of freshwater Anammox communities treating saline/brackish wastewater.

The most common way to apply Anammox for saline wastewater treatment is via salt adaptation of freshwater Anammox bacteria (FAB). To better apply this process in practice, it's essential to understand the salt adaptation process of FBA, as well as the underlying mechanisms. This study investigated the long-term salt adaptation process of a fixed-film FAB culture in three reactors (namely R1-R3), under salinities of 2, 8, and 12 NaCl g/L, correspondingly. All three reactors were under stable operation and achieved 80-90% total inorganic nitrogen removal efficiency throughout the 425-day operation period. R1 servers as a blank control, based on the clear microbial community shifts in R2 and R3, the operation period was divided into 2 phases. During Phase 1, all FAB in the three reactors belonged to Ca. Brocadia sp.. The Anammox activity (AA) and the ratio of nitrite/ammonium (NO2--N/NH4+-N) consumption in R2 and R3 decreased with the increase of salinity and did not recover to the initial levels. During Phase 2, the relative abundance of Ca. Kuenenia sp. in R2 and R3 increased from nearly 0 to about 60 and 77%, respectively. With the growth of Ca. Kuenenia sp., the AA and stoichiometry of R2 and R3 gradually recovered. AA of R2 and R3 both reached 1.0 g NH4+-N/L/day at the end of this phase, which was about 80% of that in R1. These results indicated that the salt adaptation of FAB culture was achieved by species shift from a low salt-tolerance one to a high salt-tolerance one.

Salinity effect on freshwater Anammox bacteria: Ionic stress and ion composition.

The biggest challenge to apply Anammox to treat wastewater with elevated salt content is the inhibitory effect of salinity on freshwater Anammox bacteria (FAB). Most of the research into salinity inhibition has focused on the osmotic pressure effect, while the inhibitory effect and its mechanisms induced by ion composition are poorly understood. In this study, the individual and combined effect of NaCl, KCl and Na2SO4 on FAB (>99% belonging to Ca. Brocadia genera) were systematically investigated by batch tests. The corresponding responses of mRNA abundance of three functional genes (including nitrite reductase gene (nirS), hydrazine synthase gene (hzsB) and hydrazine dehydrogenase gene (hdh)) under different salt conditions were analyzed. The results indicated that NaCl, KCl and Na2SO4 have different inhibition effects, with the 50% inhibition at 0.106, 0.096 and 0.063 M, respectively. The combined inhibition of NaCl+KCl and NaCl+Na2SO4 on FAB were both synergistic; while the combined inhibition of NaCl+KCl+Na2SO4 was additive. The responses of mRNA (of genes: nirS, hzsB and hdh) suggested NaCl inhibited the transport of ammonium; Na2SO4 inhibited both nitrite and ammonium transport; high salinity inhibited functional enzyme activity. These results suggest both ionic stress and ion composition contributed to the observed inhibition.

Changes of nitrogen-removal performance and that of the bacterial community in a mixed culture comprising freshwater and marine anammox bacteria under averaged environmental condition.

Nitrogen-removal processes using anammox bacteria are expected to achieve high-rate removal while remaining economical, and their practical applications have been investigated. However, anammox bacteria still have unfavorable characteristics for practical use, including susceptibility to a change in environmental conditions. In this study, with an aim of exploring the adaptability of mixed anammox bacteria to environmental conditions, the shift of nitrogen-removal performance and bacterial community in a mixed culture comprising freshwater anammox bacteria (FAB) and marine anammox bacteria (MAB) were investigated by a continuously stirred tank reactor (CSTR). The CSTR inoculated with the mixed anammox bacteria was operated for 180 days under an averaged condition between freshwater and marine conditions with a temperature of 27.5 °C and a synthetic medium with 15 g/L NaCl was used. Nitrogen-removal performance became stable after 114 days and more than 90% of nitrogen that was loaded into the reactor was removed in the range of nitrogen loading rate 0.07-0.42 kg N/m3/d. After operating at 0.42 kg N/m3/d for one month, a biomass sample was taken and its bacterial community was analyzed by clone-library analysis using a partial sequence of 16S rRNA. Among the clones of anammox bacteria that were made by an anammox-bacteria-specific primer, 97% of them were MAB and only 3% were FAB. These results indicate that the bacterial community including anammox bacteria was evidently changed due to environmental conditions and that the averaged condition in this study was suitable for marine bacteria rather than freshwater bacteria.