The objectives were: (a) to select the most effective lactate-utilizing Megasphaera elsdenii (Me) isolates (strains) from mixed rumen cultures (Phase 1) and (b) to determine in vivo the efficacy of the two most promising isolates from Phase 1 to control ruminal acidosis (Phase 2). In Phase 1 batch cultures of nine strains of lactate-utilizing Me were isolated and compared with the Me type strain ATCC 25940 and a membrane-filtered supernatant of a centrifuged culture (Control). Culture concentration in the incubation tubes was ca 5 x 10(5) cfu's/ml. In Phase 2 forage-fed rumen-fistulated sheep were drenched intraruminally 100 ml containing 10(11) cfu's of the promising Me isolates CH4 and CH7, following abrupt administration of 1 kg maize meal and 300g maltose per sheep, and compared with Control sheep drenched with a placebo. For Me isolates CH3, CH4 and CH7 rumen pH remained below pH = 5.0 for <3 h/24 h compared to >5 h/24 h for the type strain and the Control (p<0.001) (Phase 1). At +6 h lactic acid concentration increased to 24 mmol/l in the case of the type strain and 38.5 mmol/l for Control, compared to respectively 14 mmol/l for Me isolate CH3 and <5 mmol/l for Me isolates CH4 and CH7 (p<0.001). Lactic acid concentration for all isolates decreased to <5 mmol/l at +10 h, but for Control it progressively increased to 79 mmol/l at +24 h. In Phase 2, rumen pH for Control was <5 from 8 h to 2411 post-drenching, for Me isolate CH7 <5.5 from 4 h to 24 h, but always >5.0 (p<0.001), whereas for Me isolate CH4 rumen pH was >5.5 for the total post-drenching period. Lactic acid concentration post-drenching consistently remained <10 mmol/l for the two isolates, whereas it progressively increased to >55 mmol/l at +10 h for Control (p<0.001). Forage intake of Control sheep decreased by 16-18% from pre- to post-drenching and for Me isolate administered sheep by only 3-9% (p<0.05). It is concluded that the selected Me isolates are promising in the control of ruminal acidosis. (C) 2010 Elsevier B.V. All rights reserved.