The objective of this study was to test the effects of Pediococcus acidilactici 1 (PED) isolated from bovine cecum wall or Bacillus subtilis 2 (BAC) isolated from porcine feces as probiotics in the diet of lactating dairy cows on lactation performance and ruminal pH. To do so, 48 multiparous Holstein cows (121 ± 22 DIM) were blocked by milk yield in a randomized complete block design. All treatments consisted of a basal TMR with top-dressed supplements: (1) control (CON) with no probiotics; (2) PED fed at 1 × 10¹⁰ cfu/d; (3) BAC fed at 1 × 10¹⁰ cfu/d; and (4) basal TMR supplemented with a combination of Enterococcus faecium 3 at 1 × 10¹⁰ cfu/d and yeast (PRO). Cows were housed in a free stall barn with individual feeding gates, milked thrice a day and fed twice daily for 105 d; daily feed intake and milk yield data were averaged weekly. Eight rumen fistulated cows, 2 per treatment, were used for rumen pH measurements every 2 h during a 24-h period on d 105. Data were analyzed using a mixed model with week, treatment and their interaction as fixed effects with pre-trial milk yield as a covariate; block and cow were considered random effects. Dry matter intake was similar (P = 0.22) across treatments averaging 24.3 ± 0.76 kg/d. Milk yield was similar across treatments (P = 0.82), values were 37.2, 37.7, 36.5 and 37.9 ± 1.36 kg/d for CON, PRO, BAC and PED, respectively. Concentration of milk fat and protein were similar (P ≥ 0.69) across treatments averaging 3.63 ± 0.16% and 3.05 ± 0.06%, respectively. Overall yield of milk fat, protein and milk urea nitrogen were similar (P ≥ 0.34) across treatments and averaged 1.33 ± 0.08 kg/d, 1.11 ± 0.06 kg/d, and 12.6 ± 0.39 mg/dL, respectively. Cows consuming CON and PED tended (P = 0.12) be 28.5 kg heavier compared with the PRO and BAC. Mean daily rumen pH was similar across treatments (P = 0.29) and averaged 5.69 ± 0.05. These results demonstrate that supplementation with P. acidilactici 1 or B. subtilis 2 as probitoics for dairy cows do not have negative effects on rumen pH and that milk production and composition is maintained.