The Effect of Selenium on Nitrification and the Study of Nitrifying Bacteria Morphology
IAAAM 1991
Guy Regnier; Nancy Smith; Beverly Dixon, PhD
Department of Biological Sciences, California State University, Hayward, CA

Selenium, through the process of bioconcentration, can accumulate to reach toxic levels in the tissues of birds, fish and other wildlife. To date, little research has focused on the effect OS selenium on bacteria and in particular, on the nitrifying bacteria which provide a considerable beneficial impact on the environment. Preliminary studies in our laboratory have indicated that selenium can disrupt the nitrogen cycle by inhibiting the conversion of nitrite to nitrate by Nitrobacter spp. The effect of selenium on Nitrobacter is as of yet unknown. This break in the normal cycle allows nitrite, which is highly toxic in low levels to aquatic animals, to accumulate in the environment.

This study was undertaken to investigate the effects of selenium assimilation, by nitrifying bacteria, on the nitrogen cycle and any resultant morphological changes in cell structure.

A commercial source of nitrifying bacteria (Fritz Chemical Co., Dallas TX) was used in the study. Cultures of nitrifying bacteria were grown on Nitrosomonas medium ATCC 221, on shaker culture at 25°C. Calcium carbonate chips and Cytodex dextran microcarrier beads were provided as growing surfaces for the nitrifying bacteria. Control and experimental cultures containing varying concentrations of selenium (2.5-7.5mg/L) were tested. The chemical parameters of ammonia, nitrite, nitrate and pH were measured to determine the correlation between selenium concentration and the resultant effect on nitrification.

Bacterial samples were taken from Fritz-zyme #7 inoculated selenium cultures grown on Cytodex microcarrier dextran beads. Samples were fixed for 1 hour in 2.5% glutaraldehyde in 0.05M Na-cacolydate buffer, pH=7.0; rinsed three times in 10 minute changes of the same buffer; fixed in 1% 0s04 in 0.05M Na-cacolydate buffer prior to dehydration in a graded ethanol series.

Scanning electron microscope (SEM) samples were then dried in a critical point dryer using carbon dioxide as the transitional fluid. Specimens were then coated with 50nm of 60%-40% gold-palladium. SEM images were obtained on a Hitachi 570.

Data collected demonstrated a definite inverse relationship between the level of selenium exposed to, and the ability to convert nitrite to nitrate by Nitrobacter using light microscopy and SEM, Cytodex microcarrier dextran beads were demonstrated to provide a superior growing surface for morphology study of bacterial samples.


1.  J. G. Holt, editor, Bergey's Manual of Determinative Bacteriology, 8th edition, The Williams & Wilkins Co. Baltimore.

2.  Microcarrier Cell Culture: principles & methods, Pharmacia, Pharmacia LKB Biotechnology, Uppsala, Sweden.

3.  M.E. Hogan, H.I. Hassouna, and K.L. Klomparens, J. Electron Micro. Tech. 5; 159-169 (1987).

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Guy Regnier

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