bacterial nitrogen cycling

Role of Bacteria in Nitrogen Biogeochemical

CyclingCarlos Loyola | May 2nd, 2013

UCR, Environmental Science 301

Outline of this talk

A quick note on the Nitrogen biogeochemical cycle

Hagopian & Riley 1998. A closer look at the bacteriology of nitrification

Ruiz et al. 2003. Nitrification with high nitrite accumulation for the treatment of wastewater with high ammonia concentration.

Fdz-Polanco et al. 2000. Spatial distribution of heterotrophs and nitrifiers in a submerged biofilter for nitrification

Biogeochemical Cycle: The cycling of a chemical element or compound along the biosphere and

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The Nitrogen Cycle


Review from an aquaculture perspective

Sorry, this paper has too much

text!


Nitrogen: Proteins, nucleic acids, adenosine phosphates, nucleotides, pigments

Fish N waste: urine, feces. (In aquaculture, uneaten feed also contributes to nitrogenous waste loadin.)

Ammonia and ammonium: 60–90% of the total N excreted

Urea: 9–27% of the soluble N excreted

Hagopian paper starts here

A closer look at the bacteriology of nitrification

If food/fecal matter accumulate in water, organic compounds are proteolyzed and deaminated to ammonia.

A process called ammonification.

Further mineralization takes place by nitrification.

Both unionized ammonia and nitrite (NO2−) are toxic to fish at low concentrations.

Acute NH3 toxicity in salmonids: 0.2 mg l−1

Recommended NH3 maximum permissible level: 0.002 mg l−1 (Haywood, 1983)

Nitrite toxicity varies greatly between species and life stages, a concentration as low as 1.8 mg l−1 NO2− has been documented to be lethal to rainbow trout (Oncorhynchus mykiss) within 24 h

The lethal concentration of nitrate is 6200 mg l−1

for channel catfish (Ictalurus punctatus)


Nitrification is performed by two phylogenetically distinct groups of bacteria:

Ammonia-oxidizing bacteria (or nitrite bacteria) obtain energy by catabolizing un-ionized ammonia to nitrite

NH3+ 1.5O2 NO2− + H2O + H+ + 84 kcal mol−1

nitrite-oxidizing bacteria (or nitrate bacteria) mineralize nitrite to nitrate

NO2− + 0.5O2 NO3

− + 17.8 kcal mol−1


Nitrobacteraceae: Gram-negative chemoautotrophs, or lithotrophs.

Nitrite bacteria: Nitrosomonas, Nitrosococcus, Nitrosospira, Nitrosolobus, and Nitrosovibrio.

Nitrate bacteria: Nitrobacter, Nitrococcus, Nitrospira, and Nitrospina

Chemoautotrophic bacteria are characterized by the ability to utilize an inorganic chemical substrate (e.g. NH3, H2, Fe2+) as a source of electrons for the immobilization of inorganic carbon (i.e. CO2 (aq) or HCO3−) into biomass


This sole energy source drives carbon fixation, the assimilation of monomers into precursor metabolites, and the subsequent polymerization of building blocks and macromolecules

Chemoautotrophs are aerobic, usually employing dioxygen gas (O2) as the final or terminal electron acceptor

In contrast to photoautotrophic cyanobacteria and algae, there is no net production of oxygen.


Nitrosomonas europaea is the most abundantly cited and extensively studied nitrifier. The bacteria are short rods (0.8×1–2 mm), typically non-motile, and ubiquitous in soils (Watson, 1971). Other Nitrosomonas species have one or two subpolar flagella and inhabit freshwater and marine sediments


As autotrophs, nitrifiers are independently capable of creating the entirety of their biomass through a full complement of biosynthetic pathways

Urea and methane oxidation and the co-metabolism of a wide variety of hydrocarbons is common

Cells may be maintained under starvation conditions (i.e. ammonia or nitrite deprivation) through the low-level endogenous respiration of cytoplasmic compounds, while anabolic processes are lowered to undetectable levels


At low DO levels, ammonia-oxidizers use nitrite as an artificial electron acceptor and generate nitrous oxide (N2O) gas. Nitric oxide (NO) is produced by ammonia-oxidizers, but less sensitive to DO

Nitrifying bacteria are photoinhibited and are especially sensitive to disturbances away from optimal alkaline and mesophilic conditions

A commonly cited optimum pH for nitrifiers is 7.8

For temperature, an overall optimum of 25°C has been suggested


The maximum specific growth rate of nitrifying bacteria is uncommonly slow, owing to the low yield of their energy producing pathways and the burden of precursor formation through the incorporation of inorganic carbon

A doubling time of 7–8 h is possible under ideal conditions

Nitrifiers normally make up a very low percentage of the total microflora in soils, sediments, and naturally derived waste streams

Nitrifying biofilms are heavily overgrown by heterotrophs (pH and oxygen concentration gradient problems are exacerbated)


In natural waters nitrifiers are associated with suspended and settled particles, rather than free, unattached flotation in the water column

70 and 95% of suspended nitrifiers will cling to a fine inert media within 30 min of its introduction

If the detention time of a chemostat is not longer than the generation time of a suspended microorganism, it will rapidly be flushed out with the effluent.

Even in a flow-through system that is designed for slow growing nitrifiers, the steady-state balance may be upset by a change in the condition of the medium


End of Hagopian’s paper

Nitrification with high nitrite accumulation for the treatment of wastewater with high

ammonia concentration. Ruiz et al., 2003 Authors studied the effects of pH and dissolved

oxygen (DO) on ammonia oxidation.

Aim was to determine the conditions for saving oxygen. Aeration is a costly supply in wastewater treatment plants.

Assayed oxygen at intervals from 0.5 to 5.5 mg/L

They identified that as low as 0.7 mg/L allows the conversion of up to 98 % ammonia, accumulating up to 65% as nitrite.

Ruiz’s paper starts here

Nitrification with high nitrite accumulation for the treatment of wastewater with high ammonia

Parameters that are necessary to determine the bacteria specific growth rate μ


VSS: Volatile suspended solids, max ≅ 6.3 g/L

NLR: Nitrogen loading rate, max ≅ 3.5 kg/m3 d

Ammonia, max ≅ 780 mg N/L


Tipical removal efficiency plot (Inlet/Outlet concentrations)


Ammonia concentration at inlet (feed) 610 mg/L

DO 5.5 mg/L

pH is the dotted line


Effect of dissolved oxygen (DO): the dotted line.

NO3-out NO2

-out

NH4+

out

End of Ruiz’s paper

Spatial distribution of heterotrophs and nitrifiers in a submerged biofilter for

nitrification


nitrification

This paper describes the changes in biofilm density and specific activities of carbon, ammonia and nitrite oxidizers

Experimental setup was a nitrifying upflow biological aerated filter (UBAF)

Biochemical pathways develop as a function of the C:N ratio in the (synthetic) wastewater entering the filter.

The reactor resisted the entrance of up 200 mg COD/l without losing nitrification efficiency. This limit corresponds to a COD:NH+4 -N ratio of four.


nitrification

Organic content measured as TOC, Total Organic Carbon and COD, Chemical oxygen demand, the amount oxygen needed to chemically oxidize organic matter (a parameter describing the organic matter content)

The “entrance” zone of the filter removed 3.85 kg TOC/m3 day and 0.19 kg N/m3 day

The second zone of the filter removed 0.42 kg TOC/m3 day and 0.96 kg N/m3 day.

Spatial distribution of heterotrophs and nitrifiers

The spatial distribution of heterotrophic and nitrifying populations was quantified in terms of:

Oxygen uptake rates (OUR) or specific activities at different filter heights and

For increasing COD concentrations entering the reactor.

Specific activities of three microbial groups, ammonia oxidizers, nitrite oxidizers and heterotrophs aerobes revealed a clear microbial segregation along the filter depending on the COD concentration entering the reactor.


Head loss: Increasing pressure inside the reactor due to friction (“pressure drop” also used)


At 0.5 m heigth

At 2 m heigth


nitrification

Remarks

Critical parameters to control: DO, COD, (pH, Temp).

Configuration: DO gradient concentration along the reactor

Competition among heterotrophs/nitrifiers

Light inhibition

Questions, comments, suggestions

bacterial nitrogen cycling

Technology