[This corrects the article on p. 100 in vol. 41.].

Energy conservation in chemotrophic anaerobic bacteria.

The transformations of nitrogenous compounds by soil bacteria have beenstudied for over a century. In terms of the global fluxes between aerial and terrestrial-aquatic systems, the simplified nitrogen cycle can be envisioned as a triangle where the only biologically reversible reaction occurs between ammonium and nitrate. The reverse reactions of dinitrogen fixation or denitrification by biological means do not occur in nature. Hence, the reductive process of denitrification, defined as the reduction of nitrate or nitrite to gaseous nitrogen (usually N2), is intimately associated with the oxidative process of nitrification.

Biochemical Ecology of Nitrification and Denitrification

Publisher Summary This chapter highlights the physiology of autotrophic ammonia- and nitrite-oxidizing bacteria. Several aspects of the physiology of nitrifiers of relevance to their growth and activity in natural environments are considered. The chapter describes the basic features of the biochemistry of ammonia and nitrite oxidation and discusses the growth limiting factors and activity of these organisms. The influence of oxygen concentration, pH value, and inhibitors on their physiology is also described. Nitrification plays a central role in the nitrogen cycle of terrestrial and aquatic ecosystems, converting the most reduced form of nitrogen, NH3, to the most oxidized form, NO3. Nitrifying bacteria occupy niches in many ecosystems and compete successfully with faster and more efficiently growing organisms for oxygen, ammonia, and carbon dioxide. Nitrifiers are capable of reversing the nitrification process, carrying out denitrification and producing nitrite, ammonia, nitrous and nitric oxides, and gaseous nitrogen. Ammonia oxidizers can metabolize urea and can assimilate carbon from methane while nitrite oxidizers can grow anaerobically in the presence of organic compounds and nitrate.

Autotrophic nitrification in bacteria

Biosynthesis and metabolism of arginine in bacteria.

Reduction of nitrogenous oxides by microorganisms.

Acetate (1 to 10 mm) had no effect on the rate of nitrite oxidation or exponential growth by Nitrobacter agilis. However, acetate-1-(14)C and -2-(14)C were both assimilated by growing cultures, and acetate carbon contributed 33 to 39% of newly synthesized cell carbon. Carbon from acetate was incorporated into all of the major cell constituents, including most of the amino acids of cell protein and poly-beta-hydroxybutyrate (PHB). Cultures grown in the presence of acetate showed a significant increase in turbidity, attributable in part to protein synthesis and the accumulation of PHB in the "post-exponential phase," when the supply of nitrite was completely exhausted. Cell suspensons of N. agilis assimilated acetate in the absence of bicarbonate and even in the absence of nitrite. However, the addition of nitrite increased the rate of acetate assimilation by cell suspensions. The distribution of (14)C-acetate incorporated by cell suspensions was qualitatively similar to that found with growing cultures. Cell suspensions of N. agilis slowly oxidized acetate to CO(2). Addition of nitrite suppressed CO(2) production from acetate but increased the assimilation of acetate carbon into cell material. N. agilis contained all the enzymes of the tricarboxylic acid cycle. Growth of N. agilis in the presence of acetate did not significantly affect the levels of the enzymes of the tricarboxylic acid cycle, but did result in a 100-fold increase in the specific activity of isocitratase. In contrast, carboxydismutase was partially repressed. N. agilis was grown heterotrophically through seven transfers on a medium containing acetate and casein hydrolysate. The addition of nitrite increased the rate of heterotrophic growth. Heterotrophically grown organisms still retained their ability to grow autotrophically with nitrite. However, these organisms oxidized nitrite at a slower rate. Organisms from autotrophic and heterotrophic cultures were analyzed to determine the mean guanine plus cytosine content of their deoxyribonucleic acid; in both cases this mean was 61.2 +/- 1%. We concluded that N. agilis is not an obligate autotroph; it appears to be a facultative autotroph which resembles the novel facultative autotroph, Thiobacillus intermedius, very closely.

Acetate Assimilation by Nitrobacter agilis in Relation to Its “Obligate Autotrophy”

A new facultatively autotrophic Thiobacillus has been isolated in pure culture. The general physiological characteristics of the organism are described together with a redescription of Thiobacillus novellus. The new isolate differs from T. novellus in its ability to grow heterotrophically at faster rates and on a greater range of organic compounds. It can be transferred readily between autotrophic and heterotrophic conditions. It can grow anaerobically by nitrate respiration on a number of organic compounds, but not on thiosulfate. Some problems in the nomenclature and taxonomy of the thiobacilli are discussed with reference to the new isolate.

New Facultative Thiobacillus and a Reevaluation of the Heterotrophic Potential of Thiobacillus novellus

Hyphomicrobium species were enriched in media with methanol as sole carbon source under conditions supporting denitrification. Pure cultures of Hyphomicrobium species were isolated which denitrified vigorously with methanol. Hyphomicrobium B522, isolated by aerobic enrichment, was adapted to anaerobic growth and denitrification. Hyphomicrobium B522 and a new isolate were surveyed for anaerobic growth and denitrification on a number of simple organic compounds. Cell suspensions were tested for denitrifying activity. Nitrogen production from nitrate and nitrite and carbon dioxide production from methanol were stoichiometric.

Denitrification with Methanol: a Selective Enrichment for Hyphomicrobium Species

A new and unique obligate methylotroph was isolated from enrichment cultures with methanol as the sole source of carbon and energy. The organism grows only on methanol and methylamine and not on methane. It does not have a complex intracellular membrane system. 14C-acetate was assimilated by growing cultures and cell suspensions but was incorporated into only a limited number of cell constituents. 14C-acetate incorporation was strictly dependent on the oxidation of methanol or methylamine as a source of energy. Extracts had relatively low levels of enzymes of the tricarboxylic acid cycle, and α-ketoglutarate dehydrogenase was not detected. Comparisons were made with a facultative methylotroph isolated from the same enrichment cultures. The new obligate methylotroph contained hexose phosphate synthetase, a key enzyme in the ribose phosphate cycle of methyl metabolism.

New Obligate Methylotroph

Methylococcus capsulatus grows only on methane or methanol as its sole source of carbon and energy Some amino acids serve as nitrogen sources and are converted to keto acids which accumulate in the culture medium Cell suspensions oxidize methane, methanol, formaldehyde, and formate to carbon dioxide Other primary alcohols are oxidized only to the corresponding aldehydes Oxidation of formate by cell suspensions is more sensitive to inhibition by cyanide than is the oxidation of other one carbon compounds This is due to the cyanide sensitivity of a soluble nicotinamide adenine dinucleotide-specific formate dehydrogenase Oxidation of formaldehyde and methanol is catalyzed by a nonspecific primary alcohol dehydrogenase which is activated by ammonium ions and is independent of pyridine nucleotides Some comparisons are made with a strain of Pseudomonas methanica

Derek S. Hoare

Papers

Acetate Assimilation by Nitrobacter agilis in Relation to Its “Obligate Autotrophy”

New Facultative Thiobacillus and a Reevaluation of the Heterotrophic Potential of Thiobacillus novellus

Denitrification with Methanol: a Selective Enrichment for Hyphomicrobium Species

New Obligate Methylotroph

Physiological Studies of Methane and Methanol-Oxidizing Bacteria: Oxidation of C-1 Compounds by Methylococcus capsulatus