Background: The newly proposed methanogenic order ‘Methanomassiliicoccales' is the second largest archaeal population in the rumen, second only to the Methanobrevibacter population. However, information is limited regarding the community of this new order in the rumen.Methods: This study used real-time PCR and 454 pyrosequencing to explore the abundance and community composition of Methanomassiliicoccales in the rumen of Chinese goats fed a hay(0% grain, n = 5) or a high grain(65% grain, n = 5) diet.Results: Real-time PCR analysis showed that the relative abundance of Methanomassili coccales(% of total archaea) in the goat rumen was significantly lower in the high-grain-diet group(0.5% ± 0.2%) than that in the hay-diet group(8.2% ± 1.1%, P < 0.05). The pyrosequencing results showed that a total of 208 operational taxonomic units(OTUs) were formed from ten samples at 99% sequence identity. All the sequences were identified as Methanomassiliicoccaceae at the family level, and most of the sequences(96.82% ± 1.64%) were further classified as Group 8, 9, and 10 at the Methanomassiliicoccales genus level in each sample based on the RIM-DB database. No significant differences were observed in the number of OTUs or Chao1's, Shannon's or Pielou's evenness indexes between the hay-and high-grain-diet groups(P ≥ 0.05). PCoA analysis showed that diet altered the community of Methanomassiliicoccales.At the genus level, the relative abundances of Group 10(67.25 ± 12.76 vs. 38.13 ± 15.66, P = 0.012) and Group 4(2.07 ± 1.30 vs. 0.27 ± 0.30, P = 0.035) were significantly higher in the high-grain-diet group, while the relative abundance of Group9 was significantly higher in the hay-diet group(18.82 ± 6.20 vs. 47.14 ± 17.72, P = 0.020). At the species level, the relative abundance of Group 10 sp.(67.25 ± 12.76 vs. 38.13 ± 15.66, P = 0.012) and Group 4 sp. MpT 1(2.07 ± 1.30 vs. 0.27 ± 0.30,P = 0.035) were significantly higher in the high-grain-diet group, while the relative abundance of Group 9 sp. ISO4-G1 was signif
