International Journal of Systematic Bacteriology (1999), 49, 1083–1090 Printed in Great Britain Proposal of Virgibacillus proomii sp. nov. and emended description of Virgibacillus pantothenticus (Proom and Knight 1950) Heyndrickx et al. 1998 M. Heyndrickx, 1 † L. Lebbe, 1 K. Kersters, 1 B. Hoste, 1 R. De Wachter, 2 P. De Vos, 1 G. Forsyth 3 and N. A. Logan 3 Author for correspondence : N. A. Logan. Tel : 44 141 331 3207. Fax: 44 141 331 3208. e-mail : N.A.Logangcal.ac.uk 1 Vakgroep BFM WE10V, Laboratorium voor Microbiologie, Universiteit Gent, K. L. Ledeganckstraat 35, B-9000 Gent, Belgium 2 Department Biochemie, Universiteit Antwerpen, Universiteitsplein 1, B-2610 Antwerpen, Belgium 3 School of Biological and Biomedical Sciences, Glasgow Caledonian University, Cowcaddens Road, Glasgow G4 0BA, UK A polyphasic study of strains originally received as Bacillus (now Virgibacillus) pantothenticus, along with strains representing species belonging to Bacillus, Halobacillus and Paenibacillus, was undertaken using amplified rDNA restriction analysis (ARDRA), fatty acid methyl ester (FAME) analysis, SDS- PAGE of whole-cell proteins and routine diagnostic characters comprising 61 biochemical tests in the API system and 15 observations of vegetative cell and sporangial morphology. It revealed the presence within Virgibacillus of an as yet undescribed new species, for which the name Virgibacillus proomii is proposed ; V. proomii can be distinguished from V. pantothenticus and members of Bacillus sensu stricto, and from members of Paenibacillus and other aerobic endospore-forming bacteria, by routine phenotypic tests. The type strain of Virgibacillus proomii is LMG 12370 T . Keywords : Bacillus, amplified rDNA restriction analysis, Virgibacillus pantothenticus, Virgibacillus proomii, polyphasic taxonomy INTRODUCTION Bacillus pantothenticus was described by Proom & Knight (1950) following a nutritional analysis of mesophilic soil isolates of Bacillus species. Strains requiring pantothenic acid were isolated from different soil samples, taken from widely separated localities in Southern England. They considered them as members of a species most closely resembling Bacillus circulans but distinct from it, and subsequent studies confirmed the validity of the species (Claus & Berkeley, 1986 ; Gordon et al., 1973 ; Logan & Berkeley, 1984). Later isolations have been made from antacids (Claus & Berkeley, 1986), food, water, bile and soil. Comparisons of the 16S rRNA sequences of type strains of various Bacillus and Sporosarcina species ................................................................................................................................................. † Present address : Government Dairy Research Station, Brusselsesteen- weg 370, B-9090 Melle, Belgium. Abbreviations : ARDRA, amplified rDNA restriction analysis ; FAME, fatty acid methyl ester ; UPGMA, unweighted pair group method with arithmetic averages. The EMBL accession number for the 16S rRNA gene sequence of Virgibacillus proomii strain LMG 12370 T determined in this work is AJ012667. have indicated that B. pantothenticus lies at the periphery of rRNA group 1 of Ash et al. (1991) (Bacillus sensu stricto). Following amplified rDNA restriction analysis (ARDRA) and a polyphasic study, the new genus Virgibacillus (Heyndrickx et al., 1998) was proposed to accommodate B. pantothenticus and two related organisms which appeared to belong to an as yet undescribed new species. Polyphasic taxonomic study of further isolates received as B. pantothenticus has confirmed the existence within Virgibacillus of a second species which we propose here as Virgibacillus proomii. METHODS Strains and media. The designations of the strains, their origins and the different methods applied are shown in Table 1. Unless otherwise stated, V. pantothenticus strains were grown on Trypticase Soy agar (TSA) and the other Bacillus strains on nutrient agar with 1% (wv) glucose (NAG) at 30 C for 24–48 h. The Bacillus dipsosauri strain was grown at 37 C on TSA supplemented with 1 M KCl. The strains were checked for purity by plating and phase-contrast microscopy, and were maintained both as lyophilized cul- tures and as sporulated cultures on slopes of the appropriate above-mentioned medium containing 5 mg MnSO  4H  O 00985  1999 IUMS 1083