The rumen simulation technique (Rusitec) has been used to study the effects of phosphorus (P) deficiency on bacterial protein synthesis and chemical composition and on adenosine triphosphate (ATP) concentrations in the solid and liquid phases of fermentors. 16 g DM of a P-deficient mixed diet was put into each vessel daily and the vessels were infused with about 1 l of a P-deficient (0 mg of P) or P-supplemented (120 mg of P/l) buffer. Two vessels per treatment were used, and during a second experimental period the deficient and supplemented buffers were interchanged to determine the effects of P repletion and depletion. The proportion of bacterial N directly incorporated from dietary amino acids or peptides was greater in solid-associated bacteria (SAB) than in liquid-associated bacteria (LAB) (46 vs 19%). P deficiency increased this proportion in both bacterial populations. Protein synthesis associated with the solid phase represented about 30% of the total protein synthesis in the system. P deficiency induced a marked decrease in microbial protein synthesis in both phases. Microbial yield declined by about 5 points (g of N/kg OMF) in P-deficient conditions. ATP concentrations were greatly reduced in both phases but P deficiency had no effect on protozoal numbers. The effects of P depletion during period II were similar to those of P deficiency in period I, and P repletion showed that the effects of P deficiency were almost entirely reversible.