Rapid production of cellulase-free xylanases by solventogenic Clostridia from rumen

Xylanases without contaminating cellulases are produced by a wide variety of aerobic bacteria and fungi but anaerobic organisms producing such enzymes are rare. Clostridial species viz., Clostridium beijerinckii LU-1, C. acetobutylicum LU-1, C. butyricum and C. bifermentans LU-1 isolated from rumen produced xylanase (E.C. 3.2.1.8) of 1.5-4.2 U/ml on oat spelt xylan as sole carbon source. Celluloses absolutely did not support their growth. Among the carbon sources tested such as cellobiose, cellulose, filter paper cellulose (Whatman No. 1), methyl-nitrocellulose (MNC), AVICEL (microcrystalline cellulose); carboxymethylcellulose (CMC), starch and pectin did not support growth and enzyme activity. Among the xylans birch wood xylan and larch wood xylan did not support their growth. P-Glucosidase (E.C. 3.2.1.21), filter paper cellulase, carboxymethylcellulase (E.C. 3.2.1.4), methylnitrocellulase and avicelase (E.C. 3.2.1.74) were absent in the extracellular culture filtrate and no enzyme activity was found bound to the cells. In all these strains when grown on oat spelt xylan (1%, w/v), at pH 7 and 37 degrees C, the maximum enzyme production was observed within 21-24 It. In medium containing oat spell xylan, there was rapid increase in protein productivity, accumulation of extracellular endo-p-xylanase concomitantly with a decline of 65-82% of residual xylan, subsequent decrease in cell growth,,but the accumulated enzyme utilized the maximum amount of xylan in 24 It. C bifermentans LU-1 produced cell-bound P-xylosidase (E.C. 12.1.37) 0.145 U/ml. Endoxylanases had an optimum pH of 5 and temperature 50 degrees C (C. acetobutylicum LU-1 and C. butyricum LU-1) and (C. beijerinckii LU-1) 60 degrees C. Xylanases were stable between pH 4 and 8 and temperature 30-50 degrees C. Xylanase from C. bifermentans LU-1 had an optimum pH of 4 and temperature of 70 degrees C. The pH optima of 5 and good stability between 4 and 8 for the endoxylanases of C. beijerinckii LU-1,C. acetobutylicum LU-1 and C. butyricum LU-1 is suitable for processes where pH changes are common. (c) 2005 Elsevier Inc. All rights reserved

Isolation of obligate anaerobic rumen bacteria capable of degrading the neurotoxin beta-ODAP (beta-N-oxalyl-L-alpha,beta-diaminopropionic acid) as evaluated by a liquid chromatography/biosensor analysis system

Six pure strains of obligate anaerobes capable of degrading the toxin beta-N-oxalyl-L-alpha, beta-diaminopropionic acid (beta-ODAP) contained in grass pea (Lathyrus sativus) have been isolated from cow rumen. The new isolates were identified as Megasphaera elsdenii (five different genotypes) and Clostridium bifermentans using 16S rDNA analysis. The beta-ODAP degrading efficiency of the isolates was evaluated by measuring the amount of beta-ODAP in the growth medium, which contained beta-ODAP as the only carbon source, before and after incubation with the microbes. The method of analysis was liquid chromatography employing bioelectrochemical detection. The biosensor is based on coimmobilising two enzymes, glutamate oxidase (GlOx) and horseradish peroxidase (HRP), on the end of a spectrographic graphite electrode. beta-ODAP is oxidised by GlOx to form H2O2, which in turn is bioelectrocatalytically reduced by HRP through a mediated reaction using a polymeric mediator incorporating Os2+/3+ functionalities rapidly shuttling electrons with the electrode-giving rise to the analytical signal. On the basis of this analysis system, the new isolates are capable of utilising beta-ODAP as sole carbon source to a maximum of 90-95% within 5 days with concomitant increase in cell protein. (c) 2005 Society of Chemical Industry

Clostridium lundense sp. nov., a novel anaerobic lipolytic bacterium isolated from bovine rumen

A strictly anaerobic, mesophilic, endospore-forming, lipolytic bacterium, designated strain R1T, was isolated from bovine rumen fluid and characterized. Cells of this isolate were Gram-positive, non-motile rods that formed spherical terminal spores. The overall biochemical and physiological characteristics indicated that this strain should be placed in the genus Clostridium. The strain grew at temperatures between 25 and 47 °C (optimum, 37 °C), at pH between 5·0 and 8·5 (optimum pH 5·5–7·0) and in NaCl concentrations of 0–3 % (w/v). The isolate was not able to utilize glucose or other carbohydrates as carbon sources. The DNA G+C content was 31·2 mol%. Sequence analysis of the 16S rRNA gene of R1T revealed that it has the closest match (98 % similarity) with Clostridium tetanomorphum DSM 4474T. The highest levels of DNA–DNA relatedness of the isolate were 61·9 and 54·3 % with Clostridium pascui DSM 10365T and C. tetanomorphum DSM 4474T, respectively. Based on 16S rRNA gene sequence similarity, phylogenetic analysis, DNA G+C content, DNA–DNA hybridization data and distinct phenotypic characteristics, strain R1T (=DSM 17049T=CCUG 50446T) was classified in the genus Clostridium, as a member of a novel species, for which the name Clostridium lundense sp. nov. is proposed.Fil: Cirne, Dores G.. Lund University; SueciaFil: Delgado, Osvaldo Daniel. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Planta Piloto de Procesos Industriales Microbiológicos; ArgentinaFil: Marichamy, Sankar. Lund University; SueciaFil: Mattiasson, Bo. Lund University; Sueci