RelA-SpoT Homolog toxins pyrophosphorylate the CCA end of tRNA to inhibit protein synthesis.

RelA-SpoT Homolog (RSH) enzymes control bacterial physiology through synthesis and degradation of the nucleotide alarmone (p)ppGpp. We recently discovered multiple families of small alarmone synthetase (SAS) RSH acting as toxins of toxin-antitoxin (TA) modules, with the FaRel subfamily of toxSAS abrogating bacterial growth by producing an analog of (p)ppGpp, (pp)pApp. Here we probe the mechanism of growth arrest used by four experimentally unexplored subfamilies of toxSAS: FaRel2, PhRel, PhRel2, and CapRel. Surprisingly, all these toxins specifically inhibit protein synthesis. To do so, they transfer a pyrophosphate moiety from ATP to the tRNA 3' CCA. The modification inhibits both tRNA aminoacylation and the sensing of cellular amino acid starvation by the ribosome-associated RSH RelA. Conversely, we show that some small alarmone hydrolase (SAH) RSH enzymes can reverse the pyrophosphorylation of tRNA to counter the growth inhibition by toxSAS. Collectively, we establish RSHs as RNA-modifying enzymes.

Engineering control of bacterial cellulose production using a genetic toolkit and a new cellulose-producing strain.

Bacterial cellulose is a strong and ultrapure form of cellulose produced naturally by several species of the Acetobacteraceae Its high strength, purity, and biocompatibility make it of great interest to materials science; however, precise control of its biosynthesis has remained a challenge for biotechnology. Here we isolate a strain of Komagataeibacter rhaeticus (K. rhaeticus iGEM) that can produce cellulose at high yields, grow in low-nitrogen conditions, and is highly resistant to toxic chemicals. We achieved external control over its bacterial cellulose production through development of a modular genetic toolkit that enables rational reprogramming of the cell. To further its use as an organism for biotechnology, we sequenced its genome and demonstrate genetic circuits that enable functionalization and patterning of heterologous gene expression within the cellulose matrix. This work lays the foundations for using genetic engineering to produce cellulose-based materials, with numerous applications in basic science, materials engineering, and biotechnology.

Extracellular secretion of noncatalytic plant cell wall-binding proteins by the cellulolytic thermophile Caldicellulosiruptor bescii.

Caldicellulosiruptor bescii efficiently degrades cellulose, xylan, and native grasses at high temperatures above 70°C under anaerobic conditions. C. bescii extracellularly secretes multidomain glycoside hydrolases along with proteins of unknown function. In this study, we analyzed the C. bescii proteins that bind to the cell walls of timothy grass by using mass spectrometry, and we identified four noncatalytic plant cell wall-binding proteins (PWBPs) with high pI values (9.2 to 9.6). A search of a conserved domain database showed that these proteins possess a common domain related to solute-binding proteins. In addition, 12 genes encoding PWBP-like proteins were detected in the C. bescii genomic sequence. To analyze the binding properties of PWBPs, recombinant PWBP57 and PWBP65, expressed in Escherichia coli, were prepared. The PWBPs displayed a wide range of binding specificities: they bound to cellulose, lichenan, xylan, arabinoxylan, glucuronoxylan, mannan, glucomannan, pectin, oligosaccharides, and the cell walls of timothy grass. The proteins showed the highest binding affinity for the plant cell wall, with association constant (Ka) values of 5.2 × 10(6) to 44 × 10(6) M(-1) among the insoluble polysaccharides tested, as measured using depletion binding isotherms. Affinity gel electrophoresis demonstrated that the proteins bound to the acidic polymer pectin most strongly among the soluble polysaccharides tested. Fluorescence microscopic analysis showed that the proteins bound preferentially to the cell wall in a section of grass leaf. Binding of noncatalytic PWBPs with high pI values might be necessary for efficient utilization of polysaccharides by C. bescii at high temperatures.

Caloramator australicus sp. nov., a thermophilic, anaerobic bacterium from the Great Artesian Basin of Australia.

A strictly anaerobic, thermophilic bacterium, designated strain RC3T, was isolated from microbial mats colonizing thermal waters of a run-off channel formed by free-flowing waters from a bore well (registered no. 17263) of the Great Artesian Basin, Australia. The slightly curved rods (2.5-4.2x0.8-1.0 microm) of strain RC3T stained Gram-positive and grew optimally in tryptone-yeast extract-glucose medium at 60 degrees C (range 45-70 degrees C) and pH 7 (range pH 5-9). Strain RC3T grew poorly on yeast extract (0.2 %) but did not grow on tryptone (0.2 %) as a sole carbon source; yeast extract was required for growth on other energy sources, which included glucose, fructose, galactose, xylose, maltose, sucrose, raffinose, mannose, cellobiose, cellulose, starch, amylopectin, xylan, peptone, amyl media (Research Achievement), threonine and pyruvate but did not include arabinose, ribose, lactose, CM-cellulose, myo-inositol, mannitol, chitin, casein, formate, acetate, succinate, propionate, lactate, benzoate, glycerol, ethanol, Casamino acids, arginine, alanine, serine, glycine, glutamine, leucine, isoleucine, methionine or aspartate. The end products of glucose fermentation were ethanol and acetate. In the presence of 0.2 % yeast extract, iron(III), manganese(IV) and elemental sulfur were reduced but not sulfate, sulfite, thiosulfate, nitrate or nitrite. Iron(III) was also reduced in the presence of peptone, tryptone, amyl media, threonine and glycerol but not chitin, xylan, pectin, starch, pyruvate, acetate, benzoate, lactate, propionate, succinate, inositol, ethanol, mannitol, arginine, glutamine or serine. Strain RC3T was not able to utilize molecular hydrogen and/or carbon dioxide in the presence or absence of iron(III). In the presence of iron(III) and glycerol, increased concentrations of Fe(II) corresponded to increased cell numbers, demonstrating that strain RC3(T) was able to conserve energy to support growth from the reduction of Fe(III) to Fe(II). Chloramphenicol, streptomycin, tetracycline, penicillin and ampicillin and NaCl concentrations greater than 2 % inhibited growth. The G+C content of the DNA was 34+/-1 mol% as determined by the thermal denaturation (Tm) method. 16S rRNA gene sequence analysis indicated that strain RC3T was affiliated to Caloramator fervidus (95.8 % similarity to the type strain) and to other Caloramator species (average similarity of 91.6 %) within the phylum Firmicutes. On the basis of phylogenetic and phenotypic characteristics, it is proposed that strain RC3T should be classified in the genus Caloramator as a representative of a novel species, Caloramator australicus sp. nov. The type strain is RC3T (=JCM 1508T =KCTC 5601T).

Architecture of thermal adaptation in an Exiguobacterium sibiricum strain isolated from 3 million year old permafrost: a genome and transcriptome approach.

BACKGROUND: Many microorganisms have a wide temperature growth range and versatility to tolerate large thermal fluctuations in diverse environments, however not many have been fully explored over their entire growth temperature range through a holistic view of its physiology, genome, and transcriptome. We used Exiguobacterium sibiricum strain 255-15, a psychrotrophic bacterium from 3 million year old Siberian permafrost that grows from -5 degrees C to 39 degrees C to study its thermal adaptation. RESULTS: The E. sibiricum genome has one chromosome and two small plasmids with a total of 3,015 protein-encoding genes (CDS), and a GC content of 47.7%. The genome and transcriptome analysis along with the organism's known physiology was used to better understand its thermal adaptation. A total of 27%, 3.2%, and 5.2% of E. sibiricum CDS spotted on the DNA microarray detected differentially expressed genes in cells grown at -2.5 degrees C, 10 degrees C, and 39 degrees C, respectively, when compared to cells grown at 28 degrees C. The hypothetical and unknown genes represented 10.6%, 0.89%, and 2.3% of the CDS differentially expressed when grown at -2.5 degrees C, 10 degrees C, and 39 degrees C versus 28 degrees C, respectively. CONCLUSION: The results show that E. sibiricum is constitutively adapted to cold temperatures stressful to mesophiles since little differential gene expression was observed between 4 degrees C and 28 degrees C, but at the extremities of its Arrhenius growth profile, namely -2.5 degrees C and 39 degrees C, several physiological and metabolic adaptations associated with stress responses were observed.

In vitro effects of selected synbiotics on the human faecal microbiota composition.

Synbiotics are recognized means of modulating gut microbiota composition and activities. However, whether synbiotics are superior to prebiotics and probiotics alone in moderating the gut microbiota towards a purportedly healthy composition has not been determined. Eight selected synbiotics (short-chain fructooligosaccharides or fructooligosaccharides, each combined with one of four probiotics, Lactobacillus fermentum ME-3, Lactobacillus plantarum WCFS1, Lactobacillus paracasei 8700:2 or Bifidobacterium longum 46) were added to 24-h pH-controlled anaerobic faecal batch cultures. The prebiotic and probiotic components were also tested alone to determine their respective role within the synbiotic for modulation of the faecal microbiota. Effects upon major groups of the microbiota were evaluated using FISH. Rifampicin variant probiotic strains were used to assess probiotic levels. Synbiotic and prebiotics increased bifidobacteria and the Eubacterium rectale-Clostridium coccoides group. Lower levels of Escherichia coli were retrieved with these combinations after 5 and 10 h of fermentation. Probiotics alone had little effect upon the groups, however. Multivariate analysis revealed that the effect of synbiotics differed from the prebiotics as higher levels of Lactobacillus-Enterococcus were observed when the probiotic was stimulated by the prebiotic component. Here, the synbiotic approach was more effective than prebiotic or probiotic alone to modulate the gut microbiota.

Tindallia texcoconensis sp. nov., a new haloalkaliphilic bacterium isolated from lake Texcoco, Mexico.

A new alkaliphilic and moderately halophilic, strictly anaerobic, fermentative bacterium (strain IMP-300(T)) was isolated from a groundwater sample in the zone of the former soda lake Texcoco in Mexico. Strain IMP-300(T) was Gram-positive, non-sporulated, motile and rod-shaped. It grew within a pH range from 7.5 to 10.5, and an optimum at 9.5. The organism was obligately dependent on the presence of sodium salts. Growth showed an optimum at 35 degrees C with absence of growth above 45 degrees C. It fermented peptone and a few amino acids, preferentially arginine and ornithine, with production of acetate, propionate, and ammonium. Its fatty acid pattern was mainly composed of straight chain saturated, unsaturated, and cyclopropane fatty acids. The G + C content of genomic DNA was 40.0 mol%. Analysis of the 16S rRNA gene sequence indicated that the new isolate belongs to the genus Tindallia, in the low G + C Gram-positive phylum. Phylogenetically, strain IMP-300(T) has Tindallia californiensis, as closest relative with a 97.5% similarity level between their 16S rDNA gene sequences, but the DNA-DNA re-association value between the two DNAs was only 42.2%. On the basis of differences in genotypic, phenotypic, and phylogenetic characteristics, strain IMP-300(T) is proposed as a new species of the genus Tindallia, T. texcoconensis sp. nov. (type strain IMP-300(T ) = DSM 18041(T) = JCM 13990(T)).

Novel chemolithotrophic, thermophilic, anaerobic bacteria Thermolithobacter ferrireducens gen. nov., sp. nov. and Thermolithobacter carboxydivorans sp. nov.

Three thermophilic strains of chemolithoautotrophic Fe(III)-reducers were isolated from mixed sediment and water samples (JW/KA-1 and JW/KA-2(T): Calcite Spring, Yellowstone N.P., WY, USA; JW/JH-Fiji-2: Savusavu, Vanu Levu, Fiji). All were Gram stain positive rods (approximately 0.5 x 1.8 microm). Cells occurred singly or in V-shaped pairs, and they formed long chains in complex media. All utilized H(2) to reduce amorphous iron (III) oxide/hydroxide to magnetite at temperatures from 50 to 75 degrees C (opt. approximately 73 degrees C). Growth occurred within the pH(60C) range of 6.5-8.5 (opt. pH(60C) 7.1-7.3). Magnetite production by resting cells occurred at pH(60C) 5.5-10.3 (opt. 7.3). The iron (III) reduction rate was 1.3 mumol Fe(II) produced x h(-1) x ml(-1) in a culture with 3 x 10(7) cells, one of the highest rates reported. In the presence or absence of H(2), JW/KA-2(T) did not utilize CO. The G + C content of the genomic DNA of the type strain is 52.7 +/- 0.3 mol%. Strains JW/KA-1 and JW/KA-2(T) each contain two different 16S rRNA gene sequences. The 16S rRNA gene sequences from JW/KA-1, JW/KA-2(T), or JW/JH-Fiji-2 possessed >99% similarity to each other but also 99% similarity to the 16S rRNA gene sequence from the anaerobic, thermophilic, hydrogenogenic CO-oxidizing bacterium 'Carboxydothermus restrictus' R1. DNA-DNA hybridization between strain JW/KA-2(T) and strain R1(T) yielded 35% similarity. Physiological characteristics and the 16S rRNA gene sequence analysis indicated that the strains represent two novel species and are placed into the novel genus Thermolithobacter within the phylum 'Firmicutes'. In addition, the levels of 16S rRNA gene sequence similarity between the lineage containing the Thermolithobacter and well-established members of the three existing classes of the 'Firmicutes' is less than 85%. Therefore, Thermolithobacter is proposed to constitute the first genus within a novel class of the 'Firmicutes', Thermolithobacteria. The Fe(III)-reducing Thermolithobacter ferrireducens gen. nov., sp. nov. is designated as the type species with strain JW/KA-2(T) (ATCC 700985(T), DSM 13639(T)) as its type strain. Strain R1(T) is the type strain for the hydrogenogenic, CO-oxidizing Thermolithobacter carboxydivorans sp. nov. (DSM 7242(T), VKM 2359(T)).

Salinicoccus salsiraiae sp. nov.: a new moderately halophilic gram-positive bacterium isolated from salted skate.

Two moderately halophilic low G + C Gram-positive bacteria were isolated from a sample of salted skate (Class Chondrychthyes, Genus Raja). Phylogenetic analysis of the 16S rRNA gene sequence of strains RH1(T) and RH4 showed that these organisms represented a novel species of the genus Salinicoccus. The new isolates formed pink-red colonies and flocculated in liquid media, with optimum growth in media containing 4% NaCl and pH of about 8.0. These organisms are aerobic but reduce nitrate to nitrite under anaerobic conditions. Acid is produced from several carbohydrates. Oxidase and catalase were detected. Menaquinone 6 was the major respiratory quinone. The major fatty acids of strains RH1(T) and RH4 were 15:0 anteiso and 15:0 iso. The G + C contents of DNA were 46.2 and 46.0 mol%, respectively. The peptidoglycan was of A3alpha L-Lys-Gly(5-6) type. On the basis of the phylogenetic analyses, physiological and biochemical characteristics, we suggest that strain RH1(T) (=LMG 22840 = CIP 108576) represents a new species of the genus Salinicoccus, for which we propose the name Salinicoccus salsiraiae.

Septicemia due to Solobacterium moorei in a patient with multiple myeloma.

We report a case of bacteremia caused by Solobacterium moorei, an anaerobic, non-sporulated Gram-positive bacillus in a patient with a multiple myeloma. The source of infection was presumably related to multiple dento-alveolar abscesses. This is the first recovery of S. moorei from blood cultures.

[Evaluation of API Coryne System, version 2.0, for diphteroid gram-positive rods identification with clinical relevance]. / Evaluación del sistema API coryne, versión 2.0, para la identificación de bacilos gram-positivos difteroides de importancia clínica.

The ability of the API Coryne system, version 2.0, to identify 178 strains of gram-positive rods was evaluated. Seventy eight isolates belonged to genus Corynebacterium and one hundred to related genera, all strains were isolated from clinical samples at the Laboratory of Bacteriology, Hospital de Clínicas José de San Martin (UBA) between 1995 and 2004. The isolates were identified according to von Graevenitz and Funke's scheme. One hundred and sixty two out of 178 strains (91%) were correctly identified at genus and species level (IC95 = 85.6-94.6), in 44 of them (24.7%) additional tests were needed to final identification. Sixteen strains (9%) were not correctly identified (IC95 = 5.4-14.4); none of the 178 strains remained unidentified. The API Coryne system, version 2.0, is useful to identify the majority of Cory-nebacterium species with clinical relevance: Corynebacterium jeikeium, Corynebacterium urealyticum, Corynebacterium striatum, Corynebacterium pseudodiphtheriticum, Corynebacterium amycolatum and related species such as Arcanobacterium haemolyticum, Dermabacter hominis, Listeria monocytogenes, among others. Nevertheless for yellow-pigmented diphteroid gram-positive rods (Aureobacterium spp., Leifsonia aquatica, Microbacterium spp. and Cellulomonas spp.) and for acid fast gram-positive rods (Rhodococcus, Gordonia, Tsukamurella and Nocardia) the identification usefulness the system is limited.

Alkalibacterium iburiense sp. nov., an obligate alkaliphile that reduces an indigo dye.

Three indigo-reducing obligately alkaliphilic strains, M3(T), 41A and 41C, were isolated. The isolates grew at pH 9-12, but not at pH 7-8. They were Gram-positive, facultatively anaerobic, straight rod-shaped strains with peritrichous flagella. The isolates grew in 0-14% (w/v) NaCl, with optimum growth at 3-13%. They grew at temperatures between 10 and 45 degrees C, with optimum growth at around 30-37 degrees C. They did not hydrolyse starch or gelatin. DL-lactate was the major end-product from D-glucose. No quinones could be detected. The peptidoglycan type was A4beta, Orn-d-Asp. The major cellular fatty acids were C(16:0), C(16:1)7c and C(18:1)9c. The DNA G+C content was 42.6-43.2 mol%. Phylogenetic analysis based on 16S rRNA gene sequence data indicated that the isolates belong to the genus Alkalibacterium. DNA-DNA hybridization revealed low similarity (less than 16%) of the isolates with respect to the two closest phylogenetically related strains, Alkalibacterium olivapovliticus and Alkalibacterium psychrotolerans. On the basis of phenotypic and chemotaxonomic characteristics, phylogenetic data and DNA-DNA relatedness, the isolates merit classification as a novel species of the genus Alkalibacterium, for which the name Alkalibacterium iburiense is proposed. The type strain is M3(T) (=JCM 12662(T)=NCIMB 14024(T)).

Effects of environmental factors on leucine catabolism by Carnobacterium piscicola.

The metabolites from leucine degradation are involved in dry fermented sausage aroma. The catabolism of leucine by a strain of Carnobacterium piscicola was studied directly in the growth medium with 3H-labelled leucine to investigate the effect of five parameters: phase of growth, pH, oxygen, glucose and alpha-ketoisocaproic acid. Resting cells (RC) were also incubated with 3H-labelled leucine. The radioactive metabolites from leucine catabolism were analysed by high performance liquid chromatography (HPLC). At pH 5.4 and 7.2, the main metabolites detected were 3-methyl butanal, 3-methyl butanol and alpha-ketoisocaproic acid. At pH 6.5, the leucine catabolism was maximum and was characterised by a high production of 3-methyl butanoic acid. Leucine catabolism was most important during the exponential phase of growth. The addition of alpha-ketoisocaproic acid at 1%, glucose at levels of 0.5% to 2% and shaking of the growth medium increased leucine catabolism.

Carnobacterium viridans sp. nov., an alkaliphilic, facultative anaerobe isolated from refrigerated, vacuum-packed bologna sausage.

A facultatively anaerobic, non-spore-forming, psychrophilic, gram-positive, non-aciduric but alkaliphilic, rod-shaped bacterium (MPL-11T) was found to be responsible for green discoloration of refrigerated vacuum-packaged bologna upon opening of the package. Although Aerococcus viridans, which had been implicated earlier in causing the same problem, was also found, this is the first report of discoloration caused by an organism shown to be a species of Carnobacterium. Bacterial discoloration was caused by H2O2 production upon exposure of the meat to air. Strain MPL-11T is catalase- and oxidase-negative. It is not motile and does not reduce nitrate to nitrite or produce ammonia from arginine. It does not grow in acetate-containing broth or agar (Rogosa) or produce H2S. The peptidoglycan is of the meso-diaminopimelic acid type and it produces predominantly L(+)-lactic acid from glucose. It grows from at least 2 to 30 degrees C over a pH range from 5.5 to 9.1. Ribotyping suggested that strain MPL-11T could be a species of either Lactobacillus or Carnobacterium, but analysis using DNA sequences from the 16S rRNA gene showed conclusively that the organism belonged to the genus Carnobacterium. Since acid is not produced from amygdalin, inulin, mannitol, methyl alpha-D-glucoside or D-xylose, the organism differs from the seven described species of Carnobacterium. In addition, strain MPL-11T is the first member of the genus found that does not produce acid from ribose. It is capable of acid production/growth on galactose, glucose, fructose, mannose, N-acetylglucosamine, aesculin, cellobiose, maltose, lactose, sucrose, trehalose and tagatose. Although extremely salt tolerant, it does not grow in > or = 4% NaCl. On the basis of phenotypic and genotypic data, it is concluded that this isolate represents a separate, novel species. Accordingly, the name Carnobacterium viridans sp. nov. is proposed. The type strain is strain MPL-11T (= ATCC BAA-336T = DSM 14451T).

Factors affecting production of an antilisterial bacteriocin by Carnobacterium piscicola strain A9b in laboratory media and model fish systems.

AIMS: To investigate factors influencing bacteriocin production and bacteriocin stability of the bioprotective culture Carnobacterium piscicola strain A9b. METHODS AND RESULTS: Maximum activity was obtained in MRS7 broth (MRS adjusted to pH 7.2), with or without glucose. No bacteriocin was produced in APT broth when a low inoculum level (0.001%) was used. In contrast, inoculum level did not influence bacteriocin production in BHI and MRS7 without glucose. Bacteriocin production in APT was induced by the presence of an extracellular compound present in the sterile, filtered, cell-free supernatant fluid of a stationary-phase culture. Increasing concentrations of NaCl (2-7%) reduced bacteriocin production and maximum cell density of C. piscicola A9b when grown in cooked fish juice at 4 degrees C. CONCLUSION: Media composition, inoculum level and sodium chloride concentration affected production. SIGNIFICANCE AND IMPACT OF THE STUDY: The influence of NaCl on bacteriocin production may negate the inhibitory effect of C. piscicola A9b against Listeria monocytogenes in salty foods.

Characterization of two novel saccharolytic, anaerobic thermophiles, Thermoanaerobacterium polysaccharolyticum sp. nov. and Thermoanaerobacterium zeae sp. nov., and emendation of the genus Thermoanaerobacterium.

Two anaerobic, thermophilic, Gram-positive, non-spore forming bacteria with an array of polysaccharide-degrading enzymes were isolated from the leachate of a waste pile from a canning factory in Hoopeston, East Central Illinois, USA. The results of 16S rDNA sequence homology indicated that their closest relatives belong to the saccharolytic, thermophilic and anaerobic genera of Thermoanaerobacterium and Thermoanaerobacter. Although, the evolutionary distances between these bacteria and their closest relatives are greater than 11%, there is no defining phenotypic characteristic for the creation of a new genus. It is proposed that these bacteria should be placed in the genus Thermoanaerobacterium, which requires emendment of the genus description with regard to the reduction of thiosulfate to sulfur, because neither isolate is capable of this reduction. Thermoanaerobacterium polysaccharolyticum reduces thiosulfate to sulfide, whereas Thermoanaerobacterium zeae is unable to reduce thiosulfate. The cells of both isolates are rod-shaped and exist as single cells or sometimes in pairs. Cells are motile by means of flagella. Growth occurs between 45 and 72 degrees C, with optimum temperature of 65-68 degrees C at pH 6.8. The pH range for growth is from 4 to 8 at a temperature of 65 degrees C. Both organisms ferment glucose, arabinose, maltose, mannose, rhamnose, sucrose, trehalose, xylose, cellobiose, raffinose, melibiose and melezitose. The major end products of fermentation with glucose are ethanol and CO2, with lesser amounts of acetate, formate, lactate and hydrogen. The DNA G+C contents of Thermoanaerobacterium polysaccharolyticum sp. nov. and Thermoanaerobacterium zeae sp. nov. are 46 and 42 mol%, respectively. The type strains are KMTHCJT (= ATCC BAA-17T = DSM 13641T) and mel2T (= ATCC BAA-16T = DSM 13642T), respectively.

Bulleidia extructa gen. nov., sp. nov., isolated from the oral cavity.

Five strains of anaerobic non-sporing Gram-positive bacilli isolated from advanced periodontitis (four strains) and a dentoalveolar abscess (one strain) that did not correspond to existing species were subjected to phenotypic and genetic characterization. Following 16S rDNA sequence analysis, they were found to constitute a novel branch of the low G+C Gram-positive division of the phylogenetic tree related to Erysipelothrix rhusiopathiae and Holdemania filiformis. A new genus Bulleidia, and the species Bulleidia extructa, are proposed. Growth of B. extructa in broth media was poor but was enhanced by the addition of fructose, glucose or maltose together with Tween 80. Glucose and maltose were fermented and arginine was hydrolysed. Acetate, lactate and trace amounts of succinate were the end products of glucose fermentation. The G+C content of the DNA of the type strain is 38 mol%. The type strain of Bulleidia extructa is DSM 13220T.

Papillibacter cinnamivorans gen. nov., sp. nov., a cinnamate-transforming bacterium from a shea cake digester.

A new, strictly anaerobic, Gram-positive, non-sporulating, mesophilic bacterium, designated strain CIN1T (T=type strain) was isolated from an anaerobic digester fed with shea cake rich in tannins and aromatic compounds. Cells of strain CIN1T were rod-shaped, had characteristically pointed ends (1.3-3.0 x 0.5-0.6 microm) and occurred singly, in pairs and sometimes in chains of up to six. The pH range for growth was 6.9-8.5 and the temperature growth range was 15-40 degrees C. Optimum growth occurred with yeast extract and cinnamate at 37 degrees C and a pH of 7.5. The isolate transformed cinnamate by degrading the aliphatic side chain to produce acetate and benzoate rather than by aromatic ring cleavage or demethoxylation. The position of the methoxyl group appears to be important in the degradation of the aliphatic side chain of cinnamate; consequently, 3-methoxycinnamate and 4-methoxycinnamate, but not 2-methoxycinnamate, are transformed to produce acetate and methoxybenzoates, namely 3-methoxybenzoate and 4-methoxybenzoate, respectively. Crotonate is degraded to acetate and butyrate. The G+C content of the DNA is 56 mol%. Phylogenetic analysis of the 16S rRNA gene of strain CIN1T indicated that it was a member of the low-G+C-containing Gram-positive branch with a specific relationship to Sporobacter termitidis (sequence identity of 88%). The phylogenetic results concur with the phenotypic data which reveals that the isolate is a novel bacterium and, based on these findings, strain CIN1T (= DSM 12816T = ATCC 700879T) has been designated Papillibacter cinnamivorans gen. nov., sp. nov.

Growth kinetics of Acetobacterium sp. on methanol-formate in continuous culture.

The fermentative metabolism of Acetobacterium sp. grown on methanol-formate in continuous culture is described. The reaction stoichiometry of methanol-formate, including cells, were as follows: CH3OH + 1.13HCOOH --> 0.87CH3COOH + 0.47 cell C. Formate enhanced growth yields by approximately 60% compared with methanol-CO2-grown cultures. Comparison of yields on methanol-formate allowed calculation of an energy yield of 1.3 mol ATP per mol acetate formed during homoacetate fermentation. The magnitudes of YEG,the theoretical maximum yield of YE, and m, the maintenance coefficient, were determined by growing the organism in methanol-formate and resulted in 16.5 g cell (mol methanol catabolized)-1 and 0.674 mmol methanol catabolized (g cell)-1 h-1, respectively. It is concluded that formate might replace CO2 as a source of carboxyl donor.

Detection of a whitening fluorescent agent as an indicator of white paper biodegradation: a new approach to study the kinetics of cellulose hydrolysis by mixed cultures.

A simple and reliable method to estimate paper degradation by cellulolytic bacteria is described. This method is based on the detection in the culture medium of a fluorescent whitening agent (FWA) added to white paper during the manufacturing process. Preliminary results using a Cellulomonas strain cultivated in a liquid medium containing FWA, indicated that this component is non-toxic at a final concentration of 0.01 per thousand (v/v) and that the fluorescence decreased during the first 24 h of incubation, i.e. during exponential growth phase, suggesting an adsorption of FWA on bacterial cells. Consequently, all experiments have been performed with a liquid medium containing FWA (0.01 per thousand v/v) and white paper (8.0 g/l) as cellulose source. Mixed bacterial populations (MBPs) were prepared from refuse samples. These MBPs, which mainly consisted of bacterial rod cells, were used as inocula and fluorescence was measured after 30 h of incubation, i.e. after the stationary phase was reached. A high linear correlation (R(2) = 0.979) was found between the percentages of degraded paper (%P) deduced from residual paper weight and the fluorescence values (F) of the culture medium and the following equation between %P and F was determined: %P = 8.71x10(-5) x F. An additional experiment using a second MBP showed a strong correlation (R(2) = 0.990) between the measured %P and the %P estimated from F values, confirming the reproducibility of the method. Moreover, the time course of paper degradation by five replicate flasks from a unique MBP was set up. Paper degradation was detected 3 to 5 days after the beginning of the stationary phase. The average degradation rate between the 7th and the 11th day of incubation was 11.4% per day. Rates of paper degradation ranged from 31 to 60% after 10 days and from 77 to 88% after 3 weeks of incubation, depending on the inoculum.