Clostridium sufflavum sp nov., isolated from a methanogenic reactor treating cattle waste

A strictly anaerobic, mesophilic, cellulolytic, bacterial strain, designated CDT-1(T), was isolated from rice-straw residue from a methanogenic reactor treating waste from cattle farms. The isolation was performed using enrichment culture with filter paper as a substrate. Cells stained Gram-negative, but reacted Gram-positively in the KOH test. Cells were slightly curved rods and were motile by means of peritrichous flagella. The strain produced yellow pigment when grown on filter-paper fragments. Although spore formation was not confirmed microscopically, thermotolerant cells were produced when the strain was grown on filter paper. The optimum temperature for growth was 33 degrees C and the optimum pH was 7.4. Oxidase, catalase and nitrate-reducing activities were absent. The strain utilized xylose, fructose, glucose, cellobiose, xylooligosaccharide, cellulose (filter-paper fragments and ball-milled filter paper) and xylan. The major fermentation products were acetate, ethanol, H-2 and CO2. The major cellular fatty acids were iso-C-15:0, iso-C-14:0 and C-16:0 DMA. The cell-wall peptidoglycan contained meso-diaminopimelic acid as the diagnostic diamino acid. The genomic DNA G + C content was 40.7 mol%. On the basis of 16S rRNA gene sequence similarities, strain CDT-1(T) could be placed in cluster III of the genus Clostridium, being closely related to type strains of Clostridium hungatei (96.6% sequence similarity), Clostridium termitidis (96.2%) and Clostridium papyrosolvens (96.1%). On the basis of the cellular, physiological and phylogenetic differences between CDT-1(T) and its close relatives, this strain represents a novel species of the genus Clostridium, for which the name Clostridium sufflavum sp. nov. is proposed. The type strain is CDT-1(T) (=JCM 14807(T)=DSM 19573(T)).