Eight ruminally cannulated Nellore steers (434 ± 35 kg initial BW) were allotted to a 4x4 replicated Latin square design (21-d periods) to evaluate the effects of concentrate and virginiamycin (VM) levels in diets containing salinomycin (SL) on the ruminal population of Streptococcus bovis, Megasphaera elsdenii, Selenomonas ruminantium, and methanogenic archae. Treatments were arranged in a 2 × 2 factorial, with 2 concentrate levels (70 and 90%) and 2 VM levels (0 and 15 ppm) in the diet DM. Animals were fed once daily at 0800 h and SL was included in all diets at 13 ppm (DM basis). Ruminal fluid samples were collected at 0 and 4 h post-feeding on d 18 of each period, and DNA was extracted from samples using the Qiagen DNA stool mini kit. Ruminal bacteria and methanogenic archaea populations were assessed by real-time PCR. Statistical analyses were performed using the MIXED procedure of SAS. No effects of time of collection nor treatment x time of collection interactions were observed for any variable. The relative populations of S. bovis, M. elsdenii, and S. ruminantium were greater (P < 0.01) for steers fed the 90% concentrate diet than for animals fed less concentrate, but no effects of VM were observed for those bacteria populations (Table 1). There was a concentrate x VM interaction (P = 0.04) for the relative populations of methanogenic archaea, but within each concentrate level, no differences between VM inclusions were observed. In conclusion, the ruminal microbial populations evaluated in this study were more affected by variations in concentrate levels than by VM inclusion in the diets. Table 1. Relative populations of ruminal bacteria and methanogenic archaea in Nellore steers fed diets containing 2 concentrate (C) levels and 2 virginiamycin (VM) levels (0 and 15 ppm)

Item	70% C			90% C		SEM	P-value
0 ppm	15 ppm	0 ppm	15 ppm	C	VM	C × VM
S. bovis	1.00	0.92		5.62	8.82	0.27	<0.01	0.61	0.46
M. elsdenii	1.00	0.72		5.70	6.02	0.15	<0.01	0.76	0.67
S. ruminantium	1.00	1.22		5.13	3.07	0.11	<0.01	0.48	0.11
Methanogenic	1.00	2.06		0.44	0.23	0.08	<0.01	0.91	0.04