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New Genus, Coprococcus, Twelve New Species, and Emended Descriptions of Four Previously Described Species of Bacteria from Human Feces LILLIAN V

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New Genus, Coprococcus, Twelve New Species, and Emended Descriptions of Four Previously Described Species of Bacteria from Human Feces LILLIAN V INTERNATIONAL JOURNAL of SYSTEMATIC BACTERIOLOGY Vol. 24, No. 2 April 1974, p. 260-277 Printed in U.S.A. Copyright 0 1974 International Association of Microbiological Societies New Genus, Coprococcus, Twelve New Species, and Emended Descriptions of Four Previously Described Species of Bacteria from Human Feces LILLIAN V. HOLDEMAN and W. E. C. MOORE Anaerobe Laboratory, Virginia Polytechnic Institute and State University, Blacksburg, Virginia24061 A new genus of anaerobic cocci, Coprococcus, and 12 new species of anaerobes, Coprococcus eutactus, C. catus, C. comes, RuminococCus callidus, R. torques, Streptococcus hansenii, Bacteroides eggerthii, Eubacterium eligens, E. formicigenerans, E. hallii, Lactobacillus rogosae, and Clostridium nexile, are described. Emended descriptions and proposed neotype strains for Strepto- coccus constellatus (Prkvot) comb. nov., S. morbillorum (Prkvot) comb. nov., S. in termedius PrCvot, and Eubacterium biiorme (Eggerth) PrCvot are presented. During quantitative and qualitative studies of The moles percent guanine plus cytosine (G+C) the human fecal flora, a large number of content of the deoxyribonucleic acid (DNA) prepara- previously undescribed bacterial species was tions was determined by the thermal melting point encountered as predominant members of the (T,) method (7) by using an automatic recording flora. Of these species, those for which we have spectrophotometer (Gilford Instrument Laboratories). DNA from Escherichia coli B was included in each set nine or more isolates from six or more persons of analyses as a standard. are described here. In addition, the taxonomic position of some species of cocci is discussed, RESULTS AND DISCUSSION and for Eubacterium biforme (Eggerth) Prbvot, an emended description is presented and a Anaerobic Cocci neotype strain is designated. Coprococcus gen. nov., and new species of the A discussion of the incidence of these species genus Coprococcus as compared with other kinds of organisms of the intestinal flora is published elsewhere (8). Many strains of the anaerobic, gram-positive cocci isolated in this study did not have characteristics that would permit their inclusion in any of the described genera. These strains do MATERIALS AND METHODS not belong to any presently described genera in the family Peptococcaceae Rogosa (14). They The organisms described in this report were isolated are unlike ruminococci because they produce and characterized according to anaerobic tube culture butyric acid. They do not occur in packets, so methods and procedures previously described (5, 8, they are not members of Sarcina. They are 18). unlike Pep tococcus and Peptostreptococcus Blood agar plates (BAP) were prepared aerobically on the day of use from brain heart infusion agar because carbohydrates appear to be the main (BHIA; Difco) dehydrated base with 0.5% yeast energy source, whereas peptones are used as a extract, vitamin K-hemin solution (5), and 5% sheep nitrogen source. blood added. For this group of organisms we propose the Egg yolk agar plates (EYA) were stored in an name Coprococcus gen. nov. (Co’pro.coc’cus anaerobe jar if not used the day of preparation and Gr. n. copro feces; Gr. n. coccus berry; M.L. were streaked within 5 days of the preparation date. If masc. n. Coprococcus fecal coccus). Copro- growth did not occur, organisms were retested by coccus includes those gram-positive, anaerobic using freshly prepared medium. cocci that actively ferment carbohydrates, Plates were incubated in vented GasPak jars in an atmosphere of 90% hydrogen-10% CO, after three producing butyric and acetic acids with formic series of partial evacuation and filling with the gas or propionic and/or lactic acids. Fermentable mixture. carbohydrates are either required or are highly Production of H,S was determined in prereduced stimulatory for growth and continued subcul- SIM medium (BBL). ture. The G+C content of the DNA of the 260 VOL. 24, 1974 NEW GENUS, NEW SPECIES, AND EMENDED DESCRIPTIONS 261 strains tested is 39 to 42 mol%. The type sediment; there usually was a slight to moderate species is C. eutactus sp. nov. turbidity; after incubation for 5 days the pH Because the coprococci are gram-positive, was 4.7 to 5.0. anaerobic cocci, they are classified in the family Growth occurred equally well at 37 and 45 Peptococcaceae in the order Eubacteriales. C; growth was poor to moderate at 25 and 30 Characteristics useful in differentiating Copro- C. coccus from other genera in the family are Biochemical reactions. The characteristics of given in Table 1. Descriptions of three new 59 isolates of this species are given in Table 2. species of Coprococcus are given in Table 2 and In addition, the type strain and five other in the following discussion. strains tested fermented dextrin, reduced neu- tral red, and produced acetylmethylcarbinol (i) Coprococcus eutactus sp. nov. (Gr. adj. (AMC). The six strains tested did not ferment eu. tac’tus orderly, well disciplined [referring to adonitol, glycerol, inulin, or sorbose and there- the uniform reactions of the different strains] .) fore grow poorly, if at all, in PY media Obligately anaerobic, nonmotile, gram-positive containing these substrates. No reaction ’was cocci which usually occur in pairs. Cells may produced on EYA by five strains; one strain did decolorize easily, particularly in media contain- not grow on the surface of EYA incubated ing a fermentable carbohydrate. Cells were anaerobically. No ammonia was detected from usually round, and 0.7 to 1.3 pm in diameter; cultures in chopped meat, PY, PY-glucose, or they could be slightly elongate in peptone-yeast arginine medium. Hippurate was not hydro- extract (PY)-glucose cultures (Fig. 1). lyzed, and H2 S (SIM) was not produced. Colonies. After incubation for 5 days in Fermentation products. Fermentation pro- rumen fluid-glucose-cellobiose agar (RGCA) roll ducts (average milliequivalents per 100 ml of tubes, colonies were 0.5 to 2 mm in diameter, culture) were as follows. white or tan, lenticular, and translucent; the From PY-glucose: Formic (1.5 to 2.0), centers of the colonies were occasionally butyric (0.9), lactic (0.7), and acetic (0.5) granular or opaque. acids, sometimes with ethanol and trace On anaerobic BAP incubated for 2 days, amounts of pyruvic and succinic acids. Abun- surface colonies of the type strain were dant hydrogen was produced. punctiform, circular, entire, convex, translu- From PY-pyruvate: Formate (1.3) and ace- cent, whitish, smooth, shiny, and without tate (0.8), sometimes with trace amounts of hemolytic activity. There was no growth on lactate, butyrate, and succinate. BAP incubated in a candle jar or in an aerobic From PY: Occasionally trace amounts of atmosphere. acetic, succinic, or butyric acids. Cultural characteristics. There was poor or no Threonine was not converted to propionate. growth in PY broth without fermentable Lactate and gluconate were not used. carbohydrate. PY-glucose cultures had abun- G+C content of the DNA: 41 mol% by T, dant growth and a smooth (occasionally ropy) for the type strain. TABLE 1. Characteristics of genera of Peptococcaceae Rogosaa Peptone, a Lactic, the Butyric or other Mol% Cell major energy Carbohydrate sole major 3+ carbon vola- G+C Genus arrangement source fermented !acid product tile acids produced <T,> Peptococcus Diplococci, -+ 36-37b short chains Peptostrep- Chains +- 33-35 toccus Coprococcus Diplo cocci, sr 3942 chains Ruminococnrs Diplococci , 1 4045 chains Sarcina Packets r 29-3 1 part from Rogosa (14). Symbols: -, Negative reaction; +, positive reaction; r, required; sr, stimulatory or required; v, variable. Where two reactions are given (e.g., ‘c+-yy), the first is the more usual and the second is observed less frequently. As reported by Rogosa (1 4). TABLE 2. Reactions of new species of anaembic coccp cbprococcus Ruminococcus Strep tococcus C. eutactus C catus C. comes R. callidus R. torques hansenii Substrate or ATCC 17 ATCC 22 ATCC 18 ATCC 53 reaction 27759 (1 2/10) 27761 (14/9) 27758 (1 2/11) 27760 (27/10) - Am ygdalin a a- W- V Arab in0 se aw aw- Cello biose a W -W - a a- Esculin V - -W -W Esculin hydrolysis + + V V +- W V Fructose a a a a -W -W a Glucose a a a a a- a a Lactose a a aw a a- a a Maltose a a a a a a W- Mannitol - V W Mannose a a -W -W -W -W W- Melezit ose a a - - -a MeIibiose a a V -W a- a Raffinose a a aw a a a Ribose -W - Salicin aw a- -a -W V a- V- Sorbitol W- W-- - Starch aw a -W -W - Starch hydrolysis V W - - - - Sucrose a a a a a a Trehalose - - - V Xylose - a a -W W- Milk C- - -C - C- C C- Gas (glucose agar) 3 92 3 432 3 92 - - 3,- Reduction (resazurin) + + + + V + V Products detected Fbla F Lba Lba Lba SA SAf La2 (2,PY 9s) (2) (S,PY) (F~,PY) @YJ) (fs) =All strains were negative for fermentation of erythritol, glycogen, inositol, and rhamnose. No strain completely digested gelatin, digested milk or meat, produced indole or wtalase, reduced nitrate. Symbols: -, Negative reaction; -,no visible growth or only slight growth and negative reaction; +, positive reaction; a (carbohydrate cultures), acid (pH below 5.5); c, curd; v, variable reaction; w, weak reaction or pH between 5.5 and 6; numbers (growth and gas), amount estimated on “- to 4+” scale. Where two reactions are given (e.g., “aw”), the first was the more usual and the second was observed less frequently. Products were analyzed from carbohydrate (usually glucose or fructose) culture. Capital letters indicate 1 meq (or more) per 100 ml of culture; small letters indicate less than 1 meq/lOO ml of culture. Products in parentheses were not uniformly detected. Symbols: a, Acetic acid; b, butyric acid; c, caproic acid; f, formic acid; ib, isobutyric acid; iv, isovderic acid; 1, lactic acid; p, propionic acid; py, pyruvic acid; s, succinic acid; 2, ethanol; 4, butanol.
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