- Email: [email protected]
Transfer of Bacteroides amylophilus to a new genus Ruminobacter gen. nov., nom. rev. as Ruminobacter amylophilus comb. nov.
System. Appl. Microbiol. 8,204-207 (1986)
Transfer of Bacteroides amylophilus to a New Genus Ruminobacter gen. nov., nom. rev. as Ruminobacter amylophilus comb. nov. ERKO STACKEBRANDT 1 and HANS HIPPE 2 1 2
ln stitut fur Allgemeine Mikrobi ologie, Christian-Albrechts-Universita t, 2300 Kiel, Federal Republ ic of Germa ny Deutsche Sarnrnlung von Mikroorg ani smen, 3400 Giittingen, Federal Republic of Germany
Received April 11, 1986
Summary Result s of 165 rib osom al RN A cata loguing dem on str at e the lack of close relati on ship between Bacteroides amylopbilus AT CC 29744 and oth er members of the genu s Bacteroides. B. am ylophilus shows a remote relationship to purple sulfur bacteria and the ir non -ph ot ot rophic relat ives (gamma subdivision, as defined by Woese et al., 1984 b), while genuine memb ers of Bacteroides cluster sepa ra tely, fo rming one of the phylogenetically ancient groups of euba cter ia, Th is finding is in acco rd with differences found between the chemotaxonomic propertie s of B. amylopbilus and other Bacteroides species tested . On the basis of the se results we propose th at B. amylophilus be transferred to a new genus, Ruminobacter, as Ruminobacter amylophiius (Hamlin and Hungate 1956 ) comb. no v.
Key words: Ruminobacter - Bacteriodes - Ruminobacter amylophilus - Taxonomy
The first culture of Bacteroides amylophilus, isolated and described by Hamlin and Hungate (1956), was lost. Since then, organisms conforming to the original description of B. amylophilus have been isolated frequently from the rumen (Blackburn and Hobson, 1962; Bryant and Robinson, 1962 ; Caldwell et al., 1969; Holdeman et al., 1977 ) and Cato et al. (1978) designat ed strain H 18 isolated by Blackburn (1968 ) as the neot ype strain of B. amylopbilus. B. ampylophilus resembles other members of Bacteroides in the following cha racte rs: Gram-negative rod s, incorporati on of CO 2 into succinic acid and ferment ative production of volatile acids from carbohydrates. However, B. amylophilus differs from the majority of Bacteroides species in a number of chemota xonomic properties, e. g. fatty acid composition (Miyagawa et al., 1979) and sphingophospholipids (Kunsman and Caldwell, 1974 ; Miyagawa et al., 1978). The se differences led Miyagawa et al. (1979) and Shah and Collins (1983) to the conclu sion that B. amylophi/us should be removed from Bacteroides sensu stricto. These conclu sions are now supported by our finding that B. amylophi/us is phylogenetically unrelated to genuine members of Bacteroides. The results of 16S ribo somal RNA cataloguing reveal that B. amylophilus groups loosely with the "core" or ganisms (enterobacteria,
vibrios, oceanospirilla, Alteromonas, Photobacterium and oth ers) of the so called gamm a subdivi sion of purple bacteria and their non-phototrophic relatives (Woese et aI., 1985 b). Members of the genus Bacteroides, e. g. B. [ragilis, B. asaccharolyticus, B. distasonis, B. m elaninogenicus, B. ouatus, B. ruminicola, B. tbetaiotaomicron, B. uniformis, and B. vulgaris, form a coherent cluster within the Bacteroides - Cytophaga - Flavobacterium line of descent (Paster et aI., 1985) which is one of the main branches (" phyla") of the kingdom of eubacteria (Woese et al., 1985 a). On the basis of these results we propose that the species B. amylophilus be reclassified as the type species of a new genus for which the name Ruminobacter has been chosen. A genus named Ruminobacter with "R. paruum" as the type species was originally described by Kaars Siipesteiin (1949) to embrace Gram -negative, non -motile, an aerobic chemoheterotrophic cellulose fermenting organisms. Prevot (1966) in his "Man ual for the Classification and Determ ination of the Anaerobic Bacteria" designated the genus "Ruminobacter" as " incertae sedis" but at the same time transferred four Bacteroides species, B. amylogenes, B. ruminicola, B. succinogenes and the non-cellulolytic species B. amylophilus into this genus. The name Ruminobacter was not included in the Approved Lists of
Ruminobacter amy lopbilus comb. nov.
Bact erial Names (Skerman et al., 1980). Con sequentl y, it ha s lost its standing in nom enclature. We there fore re-use the Bame for a new genu s with Ruminobacter amylophilus comb. nov. as the type species. Description of Ruminobacter gen. no v., nom. rev. Ru.mi.no.bac't er. L. adj. ruminalis of the rum en ; M .L. masc.n. bacter equi valent of bacterium, a small rod; M.L. ma sc. n. Ruminobacter small rod of the rumen. If not otherwise indicated, the description is based on the descriptions of Hamline and Hungate (1956), Holdeman and Moore (1974), Cato et al. (1978) , and Holdeman et al. (1984 b). Gram-negative, non- spore-forming, ovalto-long rods. Chemoorganotrophs. Metabolize carbohydrates. Ferment ation products include succina te, aceta te and formate; trace amounts of lactate and eth anol may be formed. Obligatel y ana ero bic. Ca rbon diox ide requ ired which is incorporated into succinic acid. No cytochro mes. Pept idoglycan contains meso-d iarninopirnelic acid (Miyagau/a et al., 1981 ). Sphingoph osph olipid s absent (Kunsman and Caldwell, 1974 ; Miyagawa et al., 1978 ). Pred omin ant fatty acids are straight-cha in satura ted and unsaturat ed acids (Miyagawa et al., 1979). The type species of the genus shows significant 16S ribosom al RNA sequence homologies with members of the gamma subdivision of purple bacteria, e. g. enterobacteria, vibrios, oceanospirilla, legionell ae (Woese et aI., 1985 b). The mol % of the DNA (buoya nt density) is 40-42 (Reddy and Bryant, 1977 ). Th e type species is Ruminobacter amylophilus (Hamlin and Hungate, 1956) co mb. nov. Description of Ruminobacter amyJophiJus (Hamlin and Hungate 1956) comb. nov. a.my.lo'phi.lus. Gr.n . amylum sta rch; Gr. adj. philus loving ; am py lopbilus M.L. ad j. sta rch loving. Th e description is based on the type strain. After incubation for 2 da ys in PY-ma ltose broth cells are 0.9-1.2 by 1-3 urn, pleomorphic, ova l-to-Iong rods with round or tap ered ends. Swollen form s and irregular cells may occur in other media. No nmotile. Surface colonies (2 days) on rum en fluid-glucose-cello biose agar roll tube s ar e 1 mm in diameter, circular , enti re, slightly con vex, translucent , smoo th, glistening, white to tan . No gro wth occur s on the surface of plates incub ated in an aerobic jar. Colonies in deep agar are 0.8-1 mm in diameter, lent icular, ent ire or irr egula r, white and soft but yrou s. Starch broth cultures are turbid and have a final pH of 5.3-5.5. Amylase production is stimulated by Tween 80 (McWethy and Hartman, 1977 ). Carbon dioxide, ammonia and a fermentable carbohydr ate are required for gro wth. Carbon dioxide is fixed and ammonia is assimilated. Amin o acids, volatile fatty acids,
205
vita mins and haemin are not requ ired for gro wth (Blackburn, 1968 ); Miura et al., 1980; Caldwell et al. , 1973; Bryant and Robinson, 196 2). A high concent ration of sodium ion s is required for optimum growth (Caldwell et aI., 1973 ). Optimum temp erature for gro wth is about 39 °C. Hydrogen is not pr odu ced; no grow th in 20 % bile; esculin not digested; milk and meat reac tion negative. Cellulose is no t digested. Casein is not utilized but is digested when cells ar e gro wn in ammo nia containing media (Caldwell et aI., 1973). Prote ase is produced in the exponential growth phase (Blackburn, 196 8; Blackburn' and Hullah, 1974 ). Acid is produced from dext rine, glycogen, maltose and starch. No acid is produced from amygdalin, arabinose, cellobiose, esculin, fru ctose, galactose, glucose, glycerol, inositol, inulin, lactose, mannitol, raffinose, rhamnose, ribose, salicin, sucrose, trehalose, xylose. Strains are very sensitive to inhibition by heavy metals and trace elements (Forsberg, 1978). No cytochromes (Reddy and Bryant, 1977) or sphingolip ids have been detect ed (Kunsman and Caldwell, 1974; Miyagawa et al., 1978 ). Menaquinon es are absent (R. Kroppenstedt, pers. comm.). Occurs sporadically in the rum en contents of cattle but, when present , may be the pr edominant starch digester and may constitu te as much as 10% of the bacterial popul ation of the rumen. Occur s also in the ovine rumen (Bryant and Robinson, 1962; Blackburn and Hobson, 1962). Th e mol % G +C of the DN A (buoyant density) is 40-42 (Reddy and Bryant, 1977). Type strain: ATCC 29 744; DSM 136 1 (Hobson H 18). Additional taxonomic comments The results of comparative 16S rRNA cataloguing ar e, to date, the only data by which the genealogical relationship of Ruminobacter amylophilus can be evaluated. Phenot ypically the genus Ruminobacter , represented by R. amylophilus, is easily distinguished fro m other genera which contain anaero bic, non-m ot ile Gra m-negative rod s. H owe ver, if and when other species are described the generic description may have to be emended. Possible addit ion al species of Ruminobacter include or ganisms which differ in the chemotax onomic properties described by Shah and Collins (1983) for the genus Bacteroides. Of these, B. hypermegas, B. multiacidus and B. microfu sus have been reclassified in the new genera Megamonas, Mitsuokella an d Rikenella, respectively (Shah and Collins, 1982a, b; Collins et aI., 1985). Ruminobacter amylophilus differs consider ably from these or ganisms, i-n the pattern of carbohydra tes utilized, fermentation products, mol % G+C of their DNA, and in fatt y acid composition . Ruminobacter differs phenot ypically from other genera cur rentl y grouped in the family Bacteroideaceae as defined by Holdeman et al. (1984 a). Ruminobacter can be distinguished from Fusobacterium and Leptotrichia by the endp roducts of carbohydrate fermentation (Holdeman et aI., 1984 a). On the basis of metabolic p roperties Ruminobactel' can also be distinguished easily from several other genera containing Gram-negative obligate anaerobes which
206
E. Stackebrandt and H. Hippe
have been described recently, viz. Acidaminobacter (Starns and Hansen, 1984), Ilyobacter (Stieb and Schink, 1985), Pelobacter (Schink and Pfennig, 1982 a), Propiogenium (Schink and Pfenning, 1982 b), and Propionispira (Schink et aI., 1982). Ruminobacter amylophilus resembles certain Bacteroides species (B. fragilis, B. vulgatus, B. distasonis, B. ovatus, B. thetaiotaomicron, B. intermedius) in the mol% G+C content of its DNA but can be distinguished from them by the pattern of carboydrate utilization, endproducts of fermentation, absence of cytochromes and by the lack of any requirement for, or stimulation by, haemin (Macy and Probst, 1979).
References Blackburn, T. H.: Protease production by Bacteroides amylophilus H 18. J. gen. Microbiol. 53, 27-36 (1968) Blackburn, T. H., Hobson, P. N.: Further studies on the isolation of proteolytic bacteria from the sheep rumen. J. gen. MicrobioI. 29, 69-81 (1962) Blackburn, T. H., Hullah, W. A.: The cell-bound protease of Bacteroides amylophilus H 18. Canad. J. Microbiol. 20, 435-441 (1974) Bryant, M. P., Robinson, I. M.: Some nutritional characteristics of predominant culturable ruminal bacteria. ]. Bact. 84, 605-614 (1962) Caldwell, D. R., Keeney, M., van Soest, P. ].: Effects of carbon dioxide on growth and maltose fermentation by Bacteroides amylophilus. ]. Bact. 98, 668-676 (1969) Caldwell, D. R., Keeney, M., Barton, J. S., Kelley,]. F.: Sodium and other inorganic growth requirements of Bacteroides amylophilus. ]. Bact. 114, 782-789 (1973) Cato, E. P., Moore, W. E. c, Bryant, M. P.: Designation of ncotype strains for Bacteroides amylophilus Hamlin and Hungate 1956 and Bacteroides succinogenes Hungate 1950. Int.]. system. Bact. 28, 491-495 (1978) Collins, M. D., Shah, H. N., Mitsuoka, T.: Reclassification of Bacteroides microfusus (Kaneuchi and Mitsuoka) in a new genus Rikenella, as Rikenella microfusus comb. nov. System Appl. Microbiol. 6, 79-81 (1985) Forsberg, L. W.: Effects of heavy metals and other trace elements on the fermentation activity of the rumen microflora and growth of functionally important rumen bacteria. Canad. ]. Microbiol. 24, 298-306 (1978) Hamlin, L. ]., Hungate, R. E.: Culture and physiology of a starch-digesting bacterium (Bacteroides amylophilus n. sp.) from the bovine rumen. J. Bact. 72, 548-554 (1956) Holdeman, L. V., Moore, W. E. c. Bacteroides Castellani and Chalmers 1919, p. 385-405. In: Bergey's Manual of Determinative Bacterology, 8th ed. (R. C. Buchanan and N. G. Gibbon, eds.). Baltimore, Williams and Wilkins 1974 Holdeman, L. V., Cato, E. P., Moore, W. E. C. (eds.): Anaerobe Laboratory Manual, 4th ed. Blacksburg, Virginia Polytechnic Institute and State University 1977 Holdeman, L. V., Kelley, R. W., Moore, W. E. c. Bacteroidaceae, p. 602-603. In: Bergey's Manual of Determinative Bacteriology, Vol. 1 (N. R. Krieg and]. G. Holt (eds.). Baltimore-London, Williams and Wilkins 1984 a Holdeman, L. V., Kelley, R. W., Moore, W. E. c. Bacteroides Castellani and Chalmers 1919, p.604-631. In: Bergey's Manual of Determinative Bacteriology, Vol. 1. (N. R. Krieg andJ. G.
Holt (eds.), Baltimore-London, Williams and Wilkins 1984 b Kaars Siipesteiin, A.: Cellulose-decomposing bacteria from the rumen of cattle. ]. Microbial. Serol. 15, 49-52 (1949) Kunsman, J. E., Caldwell, D. R.: Comparison of the sphingolipid content of rumen Bacteroides species. Appl. Microbiol. 28, 1088-1089 (1974) Macy, J. M., Probst, I. P.: The biology of gastrointestinal bacteroides. Ann. Rev. Microbiol. 33, 561-594 (1979) McWethy, S.]., Hartman, P. A.: Purification and some properties of an extracellular alpha amylase from Bacteroides amylophilus. J. Bact. 129, 1537-1544 (1977) Miura, H., Horiguchi, M., Matsumoto, T.: Nutritional interdependence among rumen bacteria Bacteroides amylophilus, Megasphaera elsdenii, and Ruminococcus albus. Appl. Environ. Microbiol. 40, 294-300 (1980) Miyagawa, E., Azuma, R., Suto, T.: Distribution of sphingolipids in Bacteroides species. J. gen. appl. Microbiol. 24, 341-348 (1978) Miyagawa, E., Azuma, R., Suto, T.: Cellular fatty acid composition in Gram-negative obligately anaerobic rods. J. gen. appl. Microbiol. 25, 41-51 (1979) Miyagawa, E., Azuma, R., Suto, T.: Peptidoglycan composition of Gram-negative obligat anaerobic rods. J. gen. appl. MicrobioI. 27, 199-208 (1981) Paster, B. t., Ludwig, W. Weisburg, W. G., Stackebrandt, E., Reichenbach, H., Hespell, L. B., Hahn, C. M., Gibson, J., Stetter, H. 0., Woese, C. R.: A phylogenetic grouping of the bacteroides, cytophagas and certain flavobacteria. System. Appl. Microbial. 6, 34-42 (1985) Preuot, A. R.: Manual for the classification and determination of the anaerobic bacteria, p. 120-123. First American ed., translated by V. Fredette. Philadelphia, Lea and Febinger 1966 Reddy, C. A., Bryant, M. P.: Deoxyribonucleic acid base composition of certain species of the genus Bacteroides. Canad. ]. Microbial. 23, 1252-1256 (1977) Schink, B., Pfennig, N.: Fermentation of trihydrobenzenes by Pelobacter acidigallici gen. nov. sp. nov., a new strictly anaerobic non-sporeforming bacterium. Arch. Microbial. 133, 195-201 (1982 a) Schink, B., Pfennig, N.: Propiogenium modestum gen. nov. sp. nov. a new strictly anaerobic, nonsporing bacterium growing on succinate. Arch. Microbiol. 133, 209-216 (1982 b) Schink, B., Thompson, T. E., Zeikus,]. G.: Characterisation of Propionispira arboris gen. nov. sp. nov., a nitrogen-fixing anaerobe common to wetwoods of living trees. J. gen. Microbiol. 128, 2771-2780 (1982) Shah, H. N., Collins, M. D.: Reclassification of Bacteroides hypermegas (Harrison and Hansen) in a new genus Megamonas, as Megamonas bypermega comb. nov. Zbl. Bakt. Hyg. 1. Abt. Orig. C 3, 394-398 (1982 a) Shah, H. N., Collins, M. D.: Reclassification of Bacteroides multiacidus (Mitsuoka, Terada, Watanabe and Uchida) in a new genus Mitsuokella, as Mitsuokella multiacidus comb. nov. Zbl. Bakt. Hyg., L Abt. Orig. C 3, 491-494 (1982b) Shah, H. N.,· Collins, M. D.: A review: Genus Bacteroides. A chemotaxonomic perspective. ]. appl. Bact. 55, 403-416 (1983) Sherman, V. B. D., McGrowan, V., Sneath, P. H. A.: Approved Lists of Bacterial Names. Int. J. system. Bact. 30, 225-420 (1980) Stams, A. ]. M., Hansen, T. A.: Fermentation of glutamate and other compounds by Acidaminobacter hydrogenoformans gen. nov. sp. nov. an obligate anaerobe isolated from black mud. Studies with pure cultures and mixed cultures with sulfatereducing and methanogenic bacteria. Arch. Microbiol. 137, 329-337 (1984) Stieb, M., Schink, B.: A new 3-hydroxybutyrate fermenting
Ruminobacter amylophilus comb. nov. anaerobe, Ilyobaeter polytrophus, gen. nov. sp. nov., possessing various fermentation pathways. Arch. Microbiol. 140, 139-146 (1985) Woese, C. R., Stackebrandt, E., Macke, T.]., and Fox, G. E.: A phylogenetic definition of the major eubacterial taxa. System.
Appl. Microbio!' 6, 147--"151 (1985 a)
Woese, C. R., Weisburg, W. G., Hahn, C. M., Paster, B. ]., Zablen, 1. B., Lewis, B. J., Macke, T.]., Ludwig, W., Stackebrandt, E.: The phylogeny of purple bacteria: The gamma subdivision. System. Appl, Micrbio!' 6, 25-33 (1985 b)
Professor Dr. Erko Stackebrandt, Institut fiir Allgemeine Mikrobiologie der Universitat, Olshausenstr. 40-60, 0-2300 Kid
12
System. Appl. Microbial. Vol. 8/3
207
Transfer of Bacteroides amylophilus to a New Genus Ruminobacter gen. nov., nom. rev. as Ruminobacter amylophilus comb. nov. ERKO STACKEBRANDT 1 and HANS HIPPE 2 1 2
ln stitut fur Allgemeine Mikrobi ologie, Christian-Albrechts-Universita t, 2300 Kiel, Federal Republ ic of Germa ny Deutsche Sarnrnlung von Mikroorg ani smen, 3400 Giittingen, Federal Republic of Germany
Received April 11, 1986
Summary Result s of 165 rib osom al RN A cata loguing dem on str at e the lack of close relati on ship between Bacteroides amylopbilus AT CC 29744 and oth er members of the genu s Bacteroides. B. am ylophilus shows a remote relationship to purple sulfur bacteria and the ir non -ph ot ot rophic relat ives (gamma subdivision, as defined by Woese et al., 1984 b), while genuine memb ers of Bacteroides cluster sepa ra tely, fo rming one of the phylogenetically ancient groups of euba cter ia, Th is finding is in acco rd with differences found between the chemotaxonomic propertie s of B. amylopbilus and other Bacteroides species tested . On the basis of the se results we propose th at B. amylophilus be transferred to a new genus, Ruminobacter, as Ruminobacter amylophiius (Hamlin and Hungate 1956 ) comb. no v.
Key words: Ruminobacter - Bacteriodes - Ruminobacter amylophilus - Taxonomy
The first culture of Bacteroides amylophilus, isolated and described by Hamlin and Hungate (1956), was lost. Since then, organisms conforming to the original description of B. amylophilus have been isolated frequently from the rumen (Blackburn and Hobson, 1962; Bryant and Robinson, 1962 ; Caldwell et al., 1969; Holdeman et al., 1977 ) and Cato et al. (1978) designat ed strain H 18 isolated by Blackburn (1968 ) as the neot ype strain of B. amylopbilus. B. ampylophilus resembles other members of Bacteroides in the following cha racte rs: Gram-negative rod s, incorporati on of CO 2 into succinic acid and ferment ative production of volatile acids from carbohydrates. However, B. amylophilus differs from the majority of Bacteroides species in a number of chemota xonomic properties, e. g. fatty acid composition (Miyagawa et al., 1979) and sphingophospholipids (Kunsman and Caldwell, 1974 ; Miyagawa et al., 1978). The se differences led Miyagawa et al. (1979) and Shah and Collins (1983) to the conclu sion that B. amylophi/us should be removed from Bacteroides sensu stricto. These conclu sions are now supported by our finding that B. amylophi/us is phylogenetically unrelated to genuine members of Bacteroides. The results of 16S ribo somal RNA cataloguing reveal that B. amylophilus groups loosely with the "core" or ganisms (enterobacteria,
vibrios, oceanospirilla, Alteromonas, Photobacterium and oth ers) of the so called gamm a subdivi sion of purple bacteria and their non-phototrophic relatives (Woese et aI., 1985 b). Members of the genus Bacteroides, e. g. B. [ragilis, B. asaccharolyticus, B. distasonis, B. m elaninogenicus, B. ouatus, B. ruminicola, B. tbetaiotaomicron, B. uniformis, and B. vulgaris, form a coherent cluster within the Bacteroides - Cytophaga - Flavobacterium line of descent (Paster et aI., 1985) which is one of the main branches (" phyla") of the kingdom of eubacteria (Woese et al., 1985 a). On the basis of these results we propose that the species B. amylophilus be reclassified as the type species of a new genus for which the name Ruminobacter has been chosen. A genus named Ruminobacter with "R. paruum" as the type species was originally described by Kaars Siipesteiin (1949) to embrace Gram -negative, non -motile, an aerobic chemoheterotrophic cellulose fermenting organisms. Prevot (1966) in his "Man ual for the Classification and Determ ination of the Anaerobic Bacteria" designated the genus "Ruminobacter" as " incertae sedis" but at the same time transferred four Bacteroides species, B. amylogenes, B. ruminicola, B. succinogenes and the non-cellulolytic species B. amylophilus into this genus. The name Ruminobacter was not included in the Approved Lists of
Ruminobacter amy lopbilus comb. nov.
Bact erial Names (Skerman et al., 1980). Con sequentl y, it ha s lost its standing in nom enclature. We there fore re-use the Bame for a new genu s with Ruminobacter amylophilus comb. nov. as the type species. Description of Ruminobacter gen. no v., nom. rev. Ru.mi.no.bac't er. L. adj. ruminalis of the rum en ; M .L. masc.n. bacter equi valent of bacterium, a small rod; M.L. ma sc. n. Ruminobacter small rod of the rumen. If not otherwise indicated, the description is based on the descriptions of Hamline and Hungate (1956), Holdeman and Moore (1974), Cato et al. (1978) , and Holdeman et al. (1984 b). Gram-negative, non- spore-forming, ovalto-long rods. Chemoorganotrophs. Metabolize carbohydrates. Ferment ation products include succina te, aceta te and formate; trace amounts of lactate and eth anol may be formed. Obligatel y ana ero bic. Ca rbon diox ide requ ired which is incorporated into succinic acid. No cytochro mes. Pept idoglycan contains meso-d iarninopirnelic acid (Miyagau/a et al., 1981 ). Sphingoph osph olipid s absent (Kunsman and Caldwell, 1974 ; Miyagawa et al., 1978 ). Pred omin ant fatty acids are straight-cha in satura ted and unsaturat ed acids (Miyagawa et al., 1979). The type species of the genus shows significant 16S ribosom al RNA sequence homologies with members of the gamma subdivision of purple bacteria, e. g. enterobacteria, vibrios, oceanospirilla, legionell ae (Woese et aI., 1985 b). The mol % of the DNA (buoya nt density) is 40-42 (Reddy and Bryant, 1977 ). Th e type species is Ruminobacter amylophilus (Hamlin and Hungate, 1956) co mb. nov. Description of Ruminobacter amyJophiJus (Hamlin and Hungate 1956) comb. nov. a.my.lo'phi.lus. Gr.n . amylum sta rch; Gr. adj. philus loving ; am py lopbilus M.L. ad j. sta rch loving. Th e description is based on the type strain. After incubation for 2 da ys in PY-ma ltose broth cells are 0.9-1.2 by 1-3 urn, pleomorphic, ova l-to-Iong rods with round or tap ered ends. Swollen form s and irregular cells may occur in other media. No nmotile. Surface colonies (2 days) on rum en fluid-glucose-cello biose agar roll tube s ar e 1 mm in diameter, circular , enti re, slightly con vex, translucent , smoo th, glistening, white to tan . No gro wth occur s on the surface of plates incub ated in an aerobic jar. Colonies in deep agar are 0.8-1 mm in diameter, lent icular, ent ire or irr egula r, white and soft but yrou s. Starch broth cultures are turbid and have a final pH of 5.3-5.5. Amylase production is stimulated by Tween 80 (McWethy and Hartman, 1977 ). Carbon dioxide, ammonia and a fermentable carbohydr ate are required for gro wth. Carbon dioxide is fixed and ammonia is assimilated. Amin o acids, volatile fatty acids,
205
vita mins and haemin are not requ ired for gro wth (Blackburn, 1968 ); Miura et al., 1980; Caldwell et al. , 1973; Bryant and Robinson, 196 2). A high concent ration of sodium ion s is required for optimum growth (Caldwell et aI., 1973 ). Optimum temp erature for gro wth is about 39 °C. Hydrogen is not pr odu ced; no grow th in 20 % bile; esculin not digested; milk and meat reac tion negative. Cellulose is no t digested. Casein is not utilized but is digested when cells ar e gro wn in ammo nia containing media (Caldwell et aI., 1973). Prote ase is produced in the exponential growth phase (Blackburn, 196 8; Blackburn' and Hullah, 1974 ). Acid is produced from dext rine, glycogen, maltose and starch. No acid is produced from amygdalin, arabinose, cellobiose, esculin, fru ctose, galactose, glucose, glycerol, inositol, inulin, lactose, mannitol, raffinose, rhamnose, ribose, salicin, sucrose, trehalose, xylose. Strains are very sensitive to inhibition by heavy metals and trace elements (Forsberg, 1978). No cytochromes (Reddy and Bryant, 1977) or sphingolip ids have been detect ed (Kunsman and Caldwell, 1974; Miyagawa et al., 1978 ). Menaquinon es are absent (R. Kroppenstedt, pers. comm.). Occurs sporadically in the rum en contents of cattle but, when present , may be the pr edominant starch digester and may constitu te as much as 10% of the bacterial popul ation of the rumen. Occur s also in the ovine rumen (Bryant and Robinson, 1962; Blackburn and Hobson, 1962). Th e mol % G +C of the DN A (buoyant density) is 40-42 (Reddy and Bryant, 1977). Type strain: ATCC 29 744; DSM 136 1 (Hobson H 18). Additional taxonomic comments The results of comparative 16S rRNA cataloguing ar e, to date, the only data by which the genealogical relationship of Ruminobacter amylophilus can be evaluated. Phenot ypically the genus Ruminobacter , represented by R. amylophilus, is easily distinguished fro m other genera which contain anaero bic, non-m ot ile Gra m-negative rod s. H owe ver, if and when other species are described the generic description may have to be emended. Possible addit ion al species of Ruminobacter include or ganisms which differ in the chemotax onomic properties described by Shah and Collins (1983) for the genus Bacteroides. Of these, B. hypermegas, B. multiacidus and B. microfu sus have been reclassified in the new genera Megamonas, Mitsuokella an d Rikenella, respectively (Shah and Collins, 1982a, b; Collins et aI., 1985). Ruminobacter amylophilus differs consider ably from these or ganisms, i-n the pattern of carbohydra tes utilized, fermentation products, mol % G+C of their DNA, and in fatt y acid composition . Ruminobacter differs phenot ypically from other genera cur rentl y grouped in the family Bacteroideaceae as defined by Holdeman et al. (1984 a). Ruminobacter can be distinguished from Fusobacterium and Leptotrichia by the endp roducts of carbohydrate fermentation (Holdeman et aI., 1984 a). On the basis of metabolic p roperties Ruminobactel' can also be distinguished easily from several other genera containing Gram-negative obligate anaerobes which
206
E. Stackebrandt and H. Hippe
have been described recently, viz. Acidaminobacter (Starns and Hansen, 1984), Ilyobacter (Stieb and Schink, 1985), Pelobacter (Schink and Pfennig, 1982 a), Propiogenium (Schink and Pfenning, 1982 b), and Propionispira (Schink et aI., 1982). Ruminobacter amylophilus resembles certain Bacteroides species (B. fragilis, B. vulgatus, B. distasonis, B. ovatus, B. thetaiotaomicron, B. intermedius) in the mol% G+C content of its DNA but can be distinguished from them by the pattern of carboydrate utilization, endproducts of fermentation, absence of cytochromes and by the lack of any requirement for, or stimulation by, haemin (Macy and Probst, 1979).
References Blackburn, T. H.: Protease production by Bacteroides amylophilus H 18. J. gen. Microbiol. 53, 27-36 (1968) Blackburn, T. H., Hobson, P. N.: Further studies on the isolation of proteolytic bacteria from the sheep rumen. J. gen. MicrobioI. 29, 69-81 (1962) Blackburn, T. H., Hullah, W. A.: The cell-bound protease of Bacteroides amylophilus H 18. Canad. J. Microbiol. 20, 435-441 (1974) Bryant, M. P., Robinson, I. M.: Some nutritional characteristics of predominant culturable ruminal bacteria. ]. Bact. 84, 605-614 (1962) Caldwell, D. R., Keeney, M., van Soest, P. ].: Effects of carbon dioxide on growth and maltose fermentation by Bacteroides amylophilus. ]. Bact. 98, 668-676 (1969) Caldwell, D. R., Keeney, M., Barton, J. S., Kelley,]. F.: Sodium and other inorganic growth requirements of Bacteroides amylophilus. ]. Bact. 114, 782-789 (1973) Cato, E. P., Moore, W. E. c, Bryant, M. P.: Designation of ncotype strains for Bacteroides amylophilus Hamlin and Hungate 1956 and Bacteroides succinogenes Hungate 1950. Int.]. system. Bact. 28, 491-495 (1978) Collins, M. D., Shah, H. N., Mitsuoka, T.: Reclassification of Bacteroides microfusus (Kaneuchi and Mitsuoka) in a new genus Rikenella, as Rikenella microfusus comb. nov. System Appl. Microbiol. 6, 79-81 (1985) Forsberg, L. W.: Effects of heavy metals and other trace elements on the fermentation activity of the rumen microflora and growth of functionally important rumen bacteria. Canad. ]. Microbiol. 24, 298-306 (1978) Hamlin, L. ]., Hungate, R. E.: Culture and physiology of a starch-digesting bacterium (Bacteroides amylophilus n. sp.) from the bovine rumen. J. Bact. 72, 548-554 (1956) Holdeman, L. V., Moore, W. E. c. Bacteroides Castellani and Chalmers 1919, p. 385-405. In: Bergey's Manual of Determinative Bacterology, 8th ed. (R. C. Buchanan and N. G. Gibbon, eds.). Baltimore, Williams and Wilkins 1974 Holdeman, L. V., Cato, E. P., Moore, W. E. C. (eds.): Anaerobe Laboratory Manual, 4th ed. Blacksburg, Virginia Polytechnic Institute and State University 1977 Holdeman, L. V., Kelley, R. W., Moore, W. E. c. Bacteroidaceae, p. 602-603. In: Bergey's Manual of Determinative Bacteriology, Vol. 1 (N. R. Krieg and]. G. Holt (eds.). Baltimore-London, Williams and Wilkins 1984 a Holdeman, L. V., Kelley, R. W., Moore, W. E. c. Bacteroides Castellani and Chalmers 1919, p.604-631. In: Bergey's Manual of Determinative Bacteriology, Vol. 1. (N. R. Krieg andJ. G.
Holt (eds.), Baltimore-London, Williams and Wilkins 1984 b Kaars Siipesteiin, A.: Cellulose-decomposing bacteria from the rumen of cattle. ]. Microbial. Serol. 15, 49-52 (1949) Kunsman, J. E., Caldwell, D. R.: Comparison of the sphingolipid content of rumen Bacteroides species. Appl. Microbiol. 28, 1088-1089 (1974) Macy, J. M., Probst, I. P.: The biology of gastrointestinal bacteroides. Ann. Rev. Microbiol. 33, 561-594 (1979) McWethy, S.]., Hartman, P. A.: Purification and some properties of an extracellular alpha amylase from Bacteroides amylophilus. J. Bact. 129, 1537-1544 (1977) Miura, H., Horiguchi, M., Matsumoto, T.: Nutritional interdependence among rumen bacteria Bacteroides amylophilus, Megasphaera elsdenii, and Ruminococcus albus. Appl. Environ. Microbiol. 40, 294-300 (1980) Miyagawa, E., Azuma, R., Suto, T.: Distribution of sphingolipids in Bacteroides species. J. gen. appl. Microbiol. 24, 341-348 (1978) Miyagawa, E., Azuma, R., Suto, T.: Cellular fatty acid composition in Gram-negative obligately anaerobic rods. J. gen. appl. Microbiol. 25, 41-51 (1979) Miyagawa, E., Azuma, R., Suto, T.: Peptidoglycan composition of Gram-negative obligat anaerobic rods. J. gen. appl. MicrobioI. 27, 199-208 (1981) Paster, B. t., Ludwig, W. Weisburg, W. G., Stackebrandt, E., Reichenbach, H., Hespell, L. B., Hahn, C. M., Gibson, J., Stetter, H. 0., Woese, C. R.: A phylogenetic grouping of the bacteroides, cytophagas and certain flavobacteria. System. Appl. Microbial. 6, 34-42 (1985) Preuot, A. R.: Manual for the classification and determination of the anaerobic bacteria, p. 120-123. First American ed., translated by V. Fredette. Philadelphia, Lea and Febinger 1966 Reddy, C. A., Bryant, M. P.: Deoxyribonucleic acid base composition of certain species of the genus Bacteroides. Canad. ]. Microbial. 23, 1252-1256 (1977) Schink, B., Pfennig, N.: Fermentation of trihydrobenzenes by Pelobacter acidigallici gen. nov. sp. nov., a new strictly anaerobic non-sporeforming bacterium. Arch. Microbial. 133, 195-201 (1982 a) Schink, B., Pfennig, N.: Propiogenium modestum gen. nov. sp. nov. a new strictly anaerobic, nonsporing bacterium growing on succinate. Arch. Microbiol. 133, 209-216 (1982 b) Schink, B., Thompson, T. E., Zeikus,]. G.: Characterisation of Propionispira arboris gen. nov. sp. nov., a nitrogen-fixing anaerobe common to wetwoods of living trees. J. gen. Microbiol. 128, 2771-2780 (1982) Shah, H. N., Collins, M. D.: Reclassification of Bacteroides hypermegas (Harrison and Hansen) in a new genus Megamonas, as Megamonas bypermega comb. nov. Zbl. Bakt. Hyg. 1. Abt. Orig. C 3, 394-398 (1982 a) Shah, H. N., Collins, M. D.: Reclassification of Bacteroides multiacidus (Mitsuoka, Terada, Watanabe and Uchida) in a new genus Mitsuokella, as Mitsuokella multiacidus comb. nov. Zbl. Bakt. Hyg., L Abt. Orig. C 3, 491-494 (1982b) Shah, H. N.,· Collins, M. D.: A review: Genus Bacteroides. A chemotaxonomic perspective. ]. appl. Bact. 55, 403-416 (1983) Sherman, V. B. D., McGrowan, V., Sneath, P. H. A.: Approved Lists of Bacterial Names. Int. J. system. Bact. 30, 225-420 (1980) Stams, A. ]. M., Hansen, T. A.: Fermentation of glutamate and other compounds by Acidaminobacter hydrogenoformans gen. nov. sp. nov. an obligate anaerobe isolated from black mud. Studies with pure cultures and mixed cultures with sulfatereducing and methanogenic bacteria. Arch. Microbiol. 137, 329-337 (1984) Stieb, M., Schink, B.: A new 3-hydroxybutyrate fermenting
Ruminobacter amylophilus comb. nov. anaerobe, Ilyobaeter polytrophus, gen. nov. sp. nov., possessing various fermentation pathways. Arch. Microbiol. 140, 139-146 (1985) Woese, C. R., Stackebrandt, E., Macke, T.]., and Fox, G. E.: A phylogenetic definition of the major eubacterial taxa. System.
Appl. Microbio!' 6, 147--"151 (1985 a)
Woese, C. R., Weisburg, W. G., Hahn, C. M., Paster, B. ]., Zablen, 1. B., Lewis, B. J., Macke, T.]., Ludwig, W., Stackebrandt, E.: The phylogeny of purple bacteria: The gamma subdivision. System. Appl, Micrbio!' 6, 25-33 (1985 b)
Professor Dr. Erko Stackebrandt, Institut fiir Allgemeine Mikrobiologie der Universitat, Olshausenstr. 40-60, 0-2300 Kid
12
System. Appl. Microbial. Vol. 8/3
207