Prebiotic synthesis in atmospheres containing CH4, CO,and CO2

The prebiotic synthesis of organic compounds using a spark discharge on various simulated primitive earth atmospheres at 25 degrees C has been studied. Methane mixtures contained H2 + CH4 + H2O + N2 + NH3 with H2/CH4 molar ratios from 0 to 4 and pNH3 = 0.1 torr. A similar set of experiments without added NH3 was performed. The yields of amino acids (1.2 to 4.7% based on the carbon) are approximately independent of the H2/CH4 ratio and whether NH3 was present, and a wide variety of amino acids are obtained. Mixtures of H2 + CO + H2O + N2 and H2 + CO2 + H2O + N2, with and without added NH3, all gave about 2% yields of amino acids at H2/CO and H2/CO2 ratios of 2 to 4. For a H2/CO2 ratio of 0, the yield of amino acids is extremely low (10(-3)%). Glycine is almost the only amino acid produced from CO and CO2 model atmospheres. These results show that the maximum yield is about the same for the three carbon sources at high H2/carbon ratios, but that CH4 is superior at low H2/carbon ratios. In addition, CH4 gives a much greater variety of amino acids than either CO or CO2. If it is assumed that an abundance of amino acids more complex than glycine was required for the origin of life, then these results indicate the requirement for CH4 in the primitive atmosphere.     

Nutritional versatility and growth kinetics of an Aeromonas hydrophila strain isolated from drinking water.

The nutritional versatility and growth kinetics of Aeromonas hydrophila were studied to determine the nature and the growth-promoting properties of organic compounds which may serve as substrates for the growth of this organism in drinking water during treatment and distribution. As an initial screening, a total of 69 different organic compounds were tested at a concentration of 2.5 g/liter as growth substrates for 10 A. hydrophila strains. Of these strains, strain M800 attained the highest maximum colony counts in various types of drinking water and river water and was therefore used in further measurements of growth at low substrate concentrations. A mixture of 21 amino acids and a mixture of 10 long-chain fatty acids, when added to drinking water, promoted growth of strain M800 at individual compound concentrations as low as 0.1 microgram of C per liter. Mixtures of 18 carbohydrates and 18 carboxylic acids clearly enhanced growth of the organism at individual compound concentrations above 1 microgram of C per liter. Growth measurements with 63 individual substrates at a concentration of 10 micrograms of C per liter gave growth rates of greater than or equal to 0.1/h with two amino acids, nine carbohydrates, and six long-chain fatty acids. Ks values were determined for arginine (less than or equal to 0.3 micrograms of C per liter), glucose (15.9 micrograms of C per liter), acetate (11.1 micrograms of C per liter), and oleate (2.1 micrograms of C per liter). The data obtained indicate that biomass components, such as amino acids and long-chain fatty acids, can promote multiplication of aeromonads in drinking water distribution systems at concentrations as low as a few micrograms per liter.     

Nutritional versatility and growth kinetics of an Aeromonas hydrophila strain isolated from drinking water

The nutritional versatility and growth kinetics of Aeromonas hydrophila were studied to determine the nature and the growth-promoting properties of organic compounds which may serve as substrates for the growth of this organism in drinking water during treatment and distribution. As an initial screening, a total of 69 different organic compounds were tested at a concentration of 2.5 g/liter as growth substrates for 10 A. hydrophila strains. Of these strains, strain M800 attained the highest maximum colony counts in various types of drinking water and river water and was therefore used in further measurements of growth at low substrate concentrations. A mixture of 21 amino acids and a mixture of 10 long-chain fatty acids, when added to drinking water, promoted growth of strain M800 at individual compound concentrations as low as 0.1 microgram of C per liter. Mixtures of 18 carbohydrates and 18 carboxylic acids clearly enhanced growth of the organism at individual compound concentrations above 1 microgram of C per liter. Growth measurements with 63 individual substrates at a concentration of 10 micrograms of C per liter gave growth rates of greater than or equal to 0.1/h with two amino acids, nine carbohydrates, and six long-chain fatty acids. Ks values were determined for arginine (less than or equal to 0.3 micrograms of C per liter), glucose (15.9 micrograms of C per liter), acetate (11.1 micrograms of C per liter), and oleate (2.1 micrograms of C per liter). The data obtained indicate that biomass components, such as amino acids and long-chain fatty acids, can promote multiplication of aeromonads in drinking water distribution systems at concentrations as low as a few micrograms per liter.     

Dynamic aspects of vacuolar and cytosolic amino acid pools of Saccharomyces cerevisiae.

By using the Cu2+ method (Y. Ohsumi, K. Kitamoto, and Y. Anraku, J. Bacteriol. 170:2676-2682, 1988) for differential extraction of the vacuolar and cytosolic amino acid pools from yeast cells, the amino acid compositions of the two pools extracted from Saccharomyces cerevisiae cells, grown in synthetic medium supplemented with various amino acids, were determined. Histidine and lysine in the medium expanded the vacuolar pool extremely. Glutamate also accumulated in the cells, but mainly in the cytosol. The composition of amino acids in the cytosolic pool was fairly constant, in contrast to that in the vacuolar pool. Cells grown in synthetic medium supplemented with 10 mM arginine accumulated arginine in the vacuoles at a concentration of about 430 mM. This large arginine pool was metabolically active and was effectively utilized during nitrogen starvation. Arginine efflux from the vacuoles was coupled with K+ influx, with an arginine/K+ exchange ratio of 1, as judged by the initial rate. The vacuolar arginine pool was exchangeable with lysine added to the medium and was decreased by treatment of the cells with the mating pheromone, alpha-factor.     

烫伤大鼠早期口服谷氨酰胺对血浆氨基酸代谢的影响

研究烫伤早期口服谷氨酰胺 (GLN)后GLN及其相关氨基酸的代谢变化。以Wistar大鼠为烫伤模型 ,烫伤后早期口服GLN ,并以 83 5-50型氨基酸自动分析仪的生理体液法测定血浆游离氨基酸。结果烫伤后饲标准饲料(C)组血浆GLN在烫伤后 2d和 5d降低 ,与GLN代谢相关的丙氨酸和氨无显著性变化 ;GLN饲料 (G)组各时相点GLN均增加 ,1 0h和 8d增加显著 ,与GLN代谢相关的丙氨酸和血氨增加显著 ;GLN +精氨酸 (G +A)组GLN在 2d降低 ,血氨升高显著。与C组比 ,G和G +A组血浆总氨基酸、支琏氨基酸、GLN、丙氨酸、r-氨基丁酸和氨均较C组高。提示 ,烫伤后早期口服GLN能提高血GLN和与之代谢相关的氨基酸浓度。     

The Relationship between Lactobacillus mali from Cider and Lactobacillus yamanashiensis from wine

The two species, Lactobacillus mali and L. yamanashiensis have been compared using the following criteria: morphological observations and biochemical tests; amino acids of the cell wall mucopeptides; % G+C ratios and DNA homology. These tests have shown these organisms to be similar but not identical. Their similarity is, however, sufficiently great to consider them as one species. Due to priority of publication L. yamanashiensis is proposed for the specific name and culture No. 239 as the type strain. All organisms previously known as L. mali will in future be called L. yamanashiensis subsp. mali since there are enough differences at a subspecies level to distinguish the wine and cider strains.     

Adaptive responses of Salmonella enterica serovar Typhimurium DT104 and other S. Typhimurium strains and Escherichia coli O157 to low pH environments

AIMS: Cattle are a known main reservoir for acid-resistant Escherichia coli O157 and Salmonella enterica serovar Typhimurium DT104. We studied the response of S. Typhimurium DT104 to extreme low pH environments and compared their response to that of acid-resistant E. coli O157 and other S. Typhimurium phage types. METHODS AND RESULTS: Bacteria were grown in nutrient-rich medium and subsequently acid challenged at pH 2.5. We found that stationary phase cultures of various S. Typhimurium strains were able to survive a challenge for 2 h at pH 2.5. As in E. coli, the ability of S. Typhimurium to survive at pH 2.5 was shown to be dependent on the presence of amino acids, specifically arginine. The amount of proton pumping H+/ATPase, both in E. coli O157 and S. Typhimurium strains, was lower when grown at pH values <6 than after growth at pH 7.5. Cyclo fatty acid content of membranes of bacteria grown at pH values <6 was higher than that of membranes of bacteria grown at pH 7.5. CONCLUSIONS: Various S. Typhimurium strains, both DT104 and non-DT104, are able to survive for a prolonged period of time at pH 2.5. Their response to such low pH environment is seemingly similar to that of E. coli O157. SIGNIFICANCE AND IMPACT OF THE STUDY: Food-borne pathogens like S. Typhimurium DT104 and E. coli O157 form a serious threat to public health since such strains are able to survive under extreme low pH conditions as present in the human stomach. The emergence these acid-resistant strains suggests the presence of a selection barrier. The intestinal tract of ruminants fed a carbohydrate-rich diet might be such a barrier.     

The complete amino acid sequence of the low molecular weight cytosolic acid phosphatase

This paper presents the complete amino acid sequence of the low molecular weight acid phosphatase from bovine liver. This isoenzyme of the acid phosphatase family is located in the cytosol, is not inhibited by L-(+)-tartrate and fluoride ions, but is inhibited by sulfhydryl reagents. The enzyme consists of 157 amino acid residues, has an acetylated NH2 terminus, and has arginine as the COOH-terminal residue. All 8 half-cystine residues are in the free thiol form. The molecular weight calculated from the sequence is 17,953. The sequence was determined by characterizing the peptides purified by reverse-phase high performance liquid chromatography from tryptic, thermolytic, peptic, Staphylococcus aureus protease, and chymotryptic digests of the carboxymethylated protein. No sequence homologies were found with the two known acylphosphatase isoenzymes or the metalloproteins porcine uteroferrin and purple acid phosphatase from bovine spleen (both of which have acid phosphatase activity). Two half-cystines at or near the active site were identified through the reaction of the enzyme with [14C] iodoacetate in the presence or in the absence of a competitive inhibitor (i.e. inorganic phosphate). Ac-A E Q V T K S V L F V C L G N I C R S P I A E A V F R K L V T D Q N I S D N W V I D S G A V S D W N V G R S P N P R A V S C L R N H G I N T A H K A R Q V T K E D F V T F D Y I L C M D E S N L R D L N R K S N Q V K N C R A K I E L L G S Y D P Q K Q L I I E D P Y Y G N D A D F E T V Y Q Q C V R C C R A F L E K V R-OH.     

Patterns of temperature adaptation in proteins from Methanococcus and Bacillus

It has long been known that amino acid substitutions in proteins of organisms living at moderate and high temperatures (mesophiles and thermophiles, respectively) are not all symmetrical; for example, more aligned sites have lysine in mesophiles and arginine in thermophiles than have the opposite pattern. This is generally taken to indicate that certain amino acids are favored over others by selection at different temperatures. Previous comparisons of protein sequences from mesophiles and thermophiles have used relatively small numbers of sequences from a diverse array of species, meaning that only the most common amino acid substitutions could be examined and any taxon-specific patterns would be obscured. Here, we compare a large number of proteins between mesophiles and thermophiles in the archaeal genus Methanococcus and the bacterial genus Bacillus. Each genus exhibits dramatically asymmetrical substitution patterns for many pairs of amino acids. There are several pairs of amino acids for which one amino acid is favored in thermophilic Bacillus and the other is favored in thermophilic Methanococcus; this appears to result from the higher G + C content of the DNA of thermophilic Bacillus, a complication not seen in Methanococcus.     

外源精氨酸对谷氨酸棒杆菌在高葡萄糖胁迫下生长和发酵特性的影响

【目的】探究外源添加不同氨基酸和相容性溶质对谷氨酸棒杆菌(Corynebacterium glutamicum)在高糖胁迫环境下生长的影响及可能的作用机理。【方法】通过在培养基中外源添加各种氨基酸和相容性溶质,研究其对谷氨酸棒杆菌在高葡萄糖和高蔗糖胁迫下生长的影响,并分析添加精氨酸对高葡萄糖胁迫下菌株糖转运和代谢途径中关键酶转录水平的影响,以及对菌株发酵产氨基酸的影响。进一步探究了碱性氨基酸在其它棒状杆菌属中抵御高葡萄糖胁迫的潜在作用。【结果】在高葡萄糖胁迫条件下,外源添加赖氨酸、精氨酸和组氨酸后谷氨酸棒杆菌的生物量分别提高54.7%、50.0%和37.6%;而在高蔗糖胁迫条件下,添加脯氨酸和四氢嘧啶后菌株生物量增加20%以上。进一步研究表明,在高葡萄糖胁迫下,外源添加精氨酸后谷氨酸棒杆菌的葡萄糖利用速率提高约2.5倍,谷氨酸的发酵产量也增加了127.5%。此外,碱性氨基酸对其它4种棒状杆菌也具有一定的渗透保护效应。【结论】精氨酸对谷氨酸棒杆菌在高葡萄糖胁迫下具有良好的渗透保护作用,可能归因于其能促进葡萄糖的转运和代谢能力,同时发现碱性氨基酸的渗透保护效应对棒状杆菌属具有一定的普遍性。     

Experimental Studies on the Formation of Volatile Nitrogen Compounds Induced by Pseudomonas Fragi in A Synthetic Medium with Amino Acids as Source of Nitrogen

The influence of the growth of Pseudomonas fragi strain F 111 was studied in a synthetic medium. It was shown that volatile nitrogen compounds were rapidly liberated as a result of the decomposition at 5°C of arginine and histidine. A later and slower increase of TVN was observed in media, which contained lysine or urea. From the other 7 amino acids which were included in the test, no increase of TVN was observed to be caused by the strain F 11. It was shown that within the range of 0.05 to 0.8 % histidine added to the basic salt solution the amount of TVN liberated was correlated to the amount of histidine included in the media. At the TVN maximum approx. 50 % of the amino acid nitrogen of the medium was recovered as TVN. It was further shown that the liberation of TVN was correlated to the added amount of arginine and histidine included in the growth medium. The presence of lysine also influenced on the TVN maximum which was reached. In the synthetic medium to which arginine or histidine was added the growth of strain F 111 was stimulated by the presence of glucose. The acids produced by the oxidative breakdown of glucose were neutralized partly by the volatile bases produced in the decomposition of amino acids. The ability of the Pseudomonas fragi to grow in anaerobic conditions and to produce enzymes which could decompose histidine was studied. It was concluded that the anaerobic breakdown of histidine was similar to the anaerobic breakdown of arginine reported by other authors.     

Taxonomic studies on filamentous bacteria from sewage belonging to the Flavobacterium-Cytophaga complex

Ten strains of gram-negative nonmotile pigmented filamentous bacteria were isolated from activated sludge. All strains were negative in nitrate reduction, hydrolysis of gelatin, indole production and acid production from glucose (except one strain) but positive in aminopeptidase, oxidase and catalase and oxidative in the O/F-test. Two groups could be differentiated by pigmentation and acid production from saccharose (group P, pink) or maltose (group Y, yellow). Group P (6 strains) had a G+C-content in the range from 41.9 to 43.7 mol%. The menaquinone isolated was predominantly MK-7. The pattern of branched and hydroxylated fatty acids was similar to the strains in phenovar 4–5 of Oyaizu and Komagata (1981). Group Y (4 strains) had a G+C-content of 33.7–34.6 mol%; menaquinones determined were predominantly MK-6. Fatty acid patterns were similar to those of the strains of the phenovar 6–1 of Oyaizu and Komagata (1981).     

The effect of alpha-ketoglutaric acid on amino acid utilization by nonstarter Lactobacillus spp. isolated from Cheddar cheese

AIMS: To examine the effect of alpha-ketoglutaric acid (alpha-KG) on the utilization and catabolism of amino acids by strains of nonstarter lactobacilli isolated from Cheddar cheese. METHODS AND RESULTS: The effect of alpha-KG in the growth medium of nonstarter lactobacilli on amino acid metabolism, catabolite levels, peptide hydrolase and aminotransferase activities was examined. The pattern of amino acid utilization, catabolite formation and aminotransferase activity was affected by keto acid. CONCLUSIONS: Amino acid conversion into cheese aroma and flavour compounds by nonstarter lactobacilli is enhanced in the presence of alpha-ketoglutarate. SIGNIFICANCE AND IMPACT OF THE STUDY: Increasing the availability of alpha-ketoglutarate in cheese offers a possible method of reducing the maturation period by accelerating the rate of character compound formation from amino acids by the nonstarter lactobacilli.     

Chemical composition and serological analysis of the cell wall of Peptostreptococcus

Bahn, Arthur N. (Northwestern University, Chicago, Ill.), Patrick C. Y. Kung, and James A. Hayashi. Chemical composition and serological analysis of the cell wall of Peptostreptococcus. J. Bacteriol. 91:1672-1676. 1966.-Chemical and serological analyses were made of the cell wall of Peptostreptococcus to characterize taxonomically this genus of anaerobic streptococci. Cell wall hydrolysates of P. putridus strains 06 and 85, P. intermedius strains 11 and 87, and P. elsdenii strain B-159 were prepared, and the cell wall sugars were measured quantitatively by paper chromatography. Strain 85 contained only glucose, whereas strain 06 contained 93% glucose and 7% mannose. Strain 87 contained only rhamnose, and strain 11 contained approximately equal amounts of glucose and rhamnose. Strain B-159 differed from all the other strains in having a low (3.1%) content of total carbohydrate, consisting of rhamnose, galactose, and glucose. Quantitative amino acid analyses showed that the major amino compounds present in the cell wall were glutamic and aspartic acids, alanine, lysine, muramic acid, glucosamine, and galactosamine. Strains 06 and 85 possessed this complement of amino compounds, but strains 11 and 87 had relatively little aspartic acid. Strain B-159 was markedly different in having a high content of glycine and diaminopimelic acid, with only traces of lysine; it was the only strain in which teichoic acid was found. Serological analyses were made with the use of cell wall extracts as antigenic material and with homologous antisera, as well as streptococcal group antisera for groups A through S. The only strong agglutination was obtained between strain 87 antigen and group C antisera; weak agglutination was obtained with 87 against N, O, and K, and between strain 11 and groups E and F. All other antisera gave negative reactions. It is concluded that strain B-159 does not belong to the genus Peptostreptococcus, that strains 06 and 85 are members of P. putridus, and that strains 11 and 87 may be members of two different genera.     

Common Enzymes of Branched-Chain Amino Acid Catabolism in Pseudomonas putida

Two types of Pseudomonas putida PpG2 mutants which were unable to degrade branched-chain amino acids were isolated after mutagenesis and selection for ability to grow on succinate, but not valine, as a sole source of carbon. These isolates were characterized by growth on the three branched-chain amino acids (valine, isoleucine, and leucine), on the corresponding branched-chain keto acids (2-ketoisovalerate, 2-keto-3-methylvalerate, and 2-ketoisocaproate), and on other selected intermediates as carbon sources, and by their enzymatic composition. One group of mutants lost 2-ketoisovalerate-inducible branched-chain keto acid dehydrogenase that was active on all three keto acids. There was also a concomitant loss of ability to grow on all three branched-chain amino acids as well as on all three corresponding keto acids, but there was retention of ability to use subsequent intermediates in the catabolism of branched-chain amino acids. Another type of mutant showed a marked reduction in branched-chain amino acid transaminase activity and grew poorly at the expense of all three amino acids, but it utilized subsequent intermediates as carbon sources. Both the transaminase and branched-chain keto acid dehydrogenase mutants retained the ability to degrade camphor. These findings are consistent with the view that branched-chain amino acid transaminase and branched-chain keto acid dehydrogenase are common enzymes in the catabolism of valine, isoleucine, and leucine.     

Unrelatedness of Bacillus amyloliquefaciens and Bacillus subtilis

Eight strains of highly amylolytic, sporeforming bacilli (hereafter referred to as Bacillus amyloliquefaciens) were compared with respect to their taxonomic relationship to B. subtilis. The physiological-biochemical properties of these two groups of organisms showed that B. amyloliquefaciens differed from B. subtilis by their ability to grow in 10% NaCl, characteristic growth on potato plugs, increased production of alpha-amylase, and their ability to ferment lactose with the production of acid. The base compositions of the deoxyribonucleic acid (DNA) of the B. subtilis strains consistently fell in the range of 41.5 to 43.5% guanine + cytosine (G + C), whereas that of the B. amyloliquefaciens strains was in the 43.5 to 44.9% G + C range. Hybrid formation between B. subtilis W23 and B. amyloliquefaciens F DNA revealed only a 14.7 to 15.4% DNA homology between the two species. Transducing phage, SP-10, was able to propagate on B. subtilis W23 and B. amyloliquefaciens N, and would transduce B. subtilis 168 (indole(-)) and B. amyloliquefaciens N-10 (arginine(-)) to prototrophy with a frequency of 3.9 x 10(-4) and 2.4 x 10(-5) transductants per plaque-forming unit, respectively. Attempts to transduce between the two species were unsuccessful. These data show that Bacillus amyloliquefaciens is a valid species and should not be classified as a strain or variety of B. subtilis.     

The bkdR gene of Pseudomonas putida is required for expression of the bkd operon and encodes a protein related to Lrp of Escherichia coli.

Branched-chain keto acid dehydrogenase is a multienzyme complex which is required for the metabolism of the branched-chain amino acids in Pseudomonas putida. The structural genes encoding all four proteins of the bkd operon have been cloned, and their nucleotide sequences have been determined (G. Burns, K. T. Madhusudhan, K. Hatter, and J. R. Sokatch, p. 177-184 in S. Silver, A. M. Chakrabarty, B. Iglewski, and S. Kaplan [ed.], Pseudomonas: Biotransformations, Pathogenesis, and Evolving Biotechnology, American Society for Microbiology, Washington D.C., 1990). An open reading frame which encoded a protein with 36.5% amino acid identity to the leucine-responsive regulatory protein (Lrp) of Escherichia coli was found immediately upstream of the bkd operon. Chromosomal mutations affecting this gene, named bkdR, resulted in a loss of ability to use branched-chain amino acids as carbon and energy sources and failure to produce branched-chain keto acid dehydrogenase. These mutations were complemented in trans by plasmids which contained intact bkdR. Mutations affecting bkdR did not have any effect on transport of branched-chain amino acids or transamination. Therefore, the bkdR gene product must affect expression of the bkd operon and regulation must be positive. Mutations affecting bkdR could also be complemented by plasmids containing lrp of E. coli. This is the first instance of a Lrp-like protein demonstrated to regulate expression of an operon outside of E. coli.     

Amino acid limited growth of starter cultures in milk

The specific growth rates of several Streptococcus cremoris strains were 10–40% lower in milk than in other growth in media. The growth rates in milk increased when an amino acid mixture or casein was added, whereas, when milk was diluted, the specific growth rate of the streptococci decreased. This decrease could be overcome by bringing the casein concentration in the diluted milk back to the normal value (3%). This indicates that casein-hydrolysis proceeded at a rate too low for the streptococci to reach their potential maximum specific growth rates in milk so that growth in milk is essentially amino acid-limited. This was subsequently demonstrated for S. cremoris by continuous cultivation in media with low casein concentrations. At a low dilution rate casein hydrolysis was fast enough to supply the cells with enough amino acids and lactose was growth-limiting, whereas at higher dilution rates amino acids became growth-limiting. In cultures exponentially growing in milk the concentration of free amino acids was measured to determine which amino acid(s) was(were) absent and could possibly limit growth. A number of essential amino acids (leucine, methionine, glutamate and in some cases phenylalanine) were not detected and addition of these, together, stimulated the growth of S. cremoris in milk. The amino acids leucine and phenylalanine appeared to play a particularly important role in this stimulation. These two are, supposedly, the first amino acids that become limiting during growth in milk. The effect of competition for casein and amino acids by different organisms was studied in continuous cultures. At different dilution rates different strains became dominant in these mixed cultures, suggesting that differences in apparent affinity constants (K_S) for casein, leucine and glutamate existed between the strains.     

Formation of keto acids by barotolerant marine bacteria under atmospheric pressure]

Production of keto acids by washed cells of the barotolerant strains was studied under atmospheric pressure on a glucose-mineral medium in balloons made of the copolymer of ethylene with propylene. Production of keto acids in these conditions requires elevated concentrations of glucose and minimum concentrations of ammonium nitrogen. Active aeration inhibits the accumulation of keto acids by the barotolerant bacteria. The majority of the barotolerant bacteria did not form free extracellular keto acids, other bacteria liberated comparatively small amounts of these acids. The ability to accumulate keto acids was different in particular strains.     

Isolation,characterization and comparison of bacteria from swine faeces and manure storage pits

Storage of swine manure is associated with the microbiological production of a variety of odorous chemicals including ammonia, organic acids and alcohols, and sulphides. Although largely the product of microbiological activity, little is known about the microorganisms present in swine manure. In order to gain a better understanding of the types and activities of the microorganisms present, representative strains of microorganisms were isolated from faeces and stored manure slurry, identified, and physiologically characterized. For swine manure slurry samples, total anaerobe colony counts were greatest when a non-selective, habitat simulating medium containing clarified swine manure slurry was used whereas the highest counts for faecal anaerobes were obtained on rumen fluid containing medium. Faecal and slurry samples were also plated onto the appropriate medium containing the antibiotics tetracycline, erythromycin and tylosin (10 micro g ml-1, individually) and the proportional counts of organisms capable of growing in the presence of these antibiotics determined. Randomly selected isolates from the highest dilutions were identified by 16 s rDNA sequence analysis, and selected physiological characteristics were determined. The results of these examinations indicate that the predominant culturable microorganisms from these environments are obligately anaerobic, low mol percentage G + C Gram positive bacteria (Firmicutes) who are members of Clostridial, Eubacterial, and Lactobacillus/Streptococcus phylogenetic groups. Isolates similar to Sporomusa and Flexibacter/Cytophaga/Bacteroides (CFB or Bacteroidetes) groups were also obtained. Although similar overall, faecal and slurry samples differed in bacterial composition. Manure slurry samples were dominated by organisms similar to Clostridium coccoides and Enterococcus species whereas the distribution of species present in faeces appeared much broader. Whereas most of the pure cultures could be assigned to known phylogenetic groupings, few could be identified as known species. Examination of some growth and physiological characteristics of faecal and slurry isolates showed these to be primarily carbohydrate fermenters, although some were able to ferment lactate and amino acids. When the ability of manure and faecal isolates to ferment protein, peptides and amino acids was examined, a relatively small percentage of these were able to do so and most of these fermented carbohydrates in addition to the amino acid sources provided. The predominant amino acid fermenters were most closely related to C. coccoides and C. botulinum, but representatives of the Bacteroides, Staphylococcus, Enterococcus and other phylogenetic groups were also found. The results reported here are compared with those obtained from clone libraries prepared from the same environmental samples.