To determine the effect of feeding propionibacteria on metabolic indicators during lactation, multiparous and primiparous Holstein cows were fed one of three dietary treatments in a 2×3 factorial design from 2 weeks prepartum to 30 weeks post partum: (1) Control (primiparous n=5, multiparous n=8) fed a total mixed ration (TMR); (2) high-dose group (primiparous n=6, multiparous n=5) fed TMR plus 6×1011cfu/head daily (high-dose P169) of propionibacterium strain P169; or (3) low-dose group (primiparous n=8, multiparous n=6) fed TMR plus 6×1010 cfu/head daily (low-dose P169) of P169. Blood samples were collected weekly and analysed for plasma concentrations of glucose, insulin, insulin-like growth factor-I (IGF-I), leptin, nonesterified fatty acids (NEFA) and cholesterol. Between weeks 25 and 30, all groups received bovine somatotropin (bST) every 2 weeks. Low-dose P169 multiparous cows had lower (P<0·05) plasma insulin and glucose concentrations than high-dose P169 multiparous cows, whereas high-dose P169 primiparous cows had lower glucose but greater insulin concentartions than low-dose P169 primiparous cows (P<0·05). Plasma insulin[ratio ]glucose molar ratios were 13–18% lower (P<0·05) in low-dose P169 cows than in control or high-dose P169 cows. Plasma IGF-I, NEFA and leptin levels did not differ among diet groups between weeks 1 and 25. Low-dose P169 multiparous cows had 25% greater plasma cholesterol levels than high-dose P169 and control multiparous cows, but cholesterol levels in primiparous cows did not differ. During bST treatment, high-dose P169 multiparous cows and low-dose P169 primiparous cows had lower IGF-I levels than their respective controls and, regardless of parity, high-dose P169 cows had greater NEFA than control cows. Although supplemental feeding of P169 altered plasma hormones and metabolites, the particular effects were dependent on dose of P169 and parity of cows.