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

Abstract

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 6C 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 CO 2 . 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 (5JCM 14807 T 5DSM 19573 T ).

Development of anaerobic bacterial community consisted of diverse clostridial species during biological soil disinfestation amended with plant biomass

“…Diluted nutrient broth (DNB) agar [0.1 g each of meat extract and peptone, 0.05 g of sodium chloride (NaCl), and 15 g of agar per L] was used for enumeration of aerobic bacteria. For enumeration of anaerobic bacteria, 1/10 PY4S medium that contained one-tenth amounts of peptone (Trypticase, BBL) and yeast extract in PY4S agar (Nishiyama et al 2009) was used with oxygen-free 95% nitrogen (N 2 )/5% carbon dioxide (CO 2 ) mixed gas as a headspace. All the petri dishes and roll tubes inoculated were incubated at 30 C for two weeks and each viable count (CFU g À1 ¼ colony forming units per g of dry soil) was determined.…”

“…Both cellulose and xylan are major components of plant biomass and the class Clostridia includes many anaerobic species decomposing these compounds (Bergquist et al 1999;Carere et al 2008;Rainey et al 2009). Thus, these slowly decomposable plant materials might support growth of some relatively slow-growing clostridial species, including cellulolytic or xylanolytic species (e.g., Clostridium sufflavum) (Nishiyama et al 2009), which represented new and different profiles at the later stages (15-18 days). In fact, the phylogenetic composition of major groups assigned to the Clostridia in both biomasstreated libraries largely changed at 18 d for both biomass-treated soils.…”

Bacteroides graminisolvens sp. nov., a xylanolytic anaerobe isolated from a methanogenic reactor treating cattle waste

“…In this study to enrich for xylanolytic bacteria, the peptone and yeast extract concentrations in the medium were reduced to onetenth of those in the normal PY medium. In our experience, higher concentrations of these components in medium used for the enrichment of xylan-or cellulosedecomposing bacteria usually enhanced the growth of fastgrowing proteolytic or aminolytic bacteria, and these bacteria usually outcompeted polysaccharide-decomposing bacteria (Nishiyama et al, 2009). The use of reduced concentrations of peptone and yeast extract should suppress the growth of non-xylanolytic bacteria compared with that produced in the normal PY medium, and thus xylan-decomposing Bacteroides species should be enriched successfully.…”

“…PY medium supplemented with 0.25 g each of glucose, cellobiose, maltose and soluble starch l 21 as well as 15 g agar l 21 (Difco) was designated PY4S agar and used for maintenance of the strain in agar slants. For enrichment culture and the isolation of xylanolytic bacteria, the concentrations of both peptone and yeast extract in the basal medium were decreased to one-tenth of those in PY medium (1/10PY medium) (Nishiyama et al, 2009 (Holdeman et al, 1977) (PYH medium) and 10 g glucose l 21 (PYHG medium) was used for cultivation of the strain for various physiological tests and chemotaxonomic analyses of the cells, unless otherwise stated (Ueki et al, 2006b). In addition to haemin, a Bvitamin mixture (10 ml l…”

Descriptions of Anaerotaenia torta gen. nov., sp. nov. and Anaerocolumna cellulosilytica gen. nov., sp. nov. isolated from a methanogenic reactor of cattle waste and reclassification of Clostridium aminovalericum, Clostridium jejuense and Clostridium xylanovorans as Anaerocolumna species

“…Strains FH052 T and SN021 T were isolated by the anaerobic roll-tube method (Holdeman et al, 1977) from a methanogenic reactor treating waste collected from cattle farms (housing up to 1000 cattle in total) in Betsukai-machi, Hokkaido, Japan, as described previously (Nishiyama et al, 2009;Ueki et al, 2009Ueki et al, , 2014Ueki et al, , 2015. Strain FH052 T was from digester fluid without plant residue or other solid material and strain SN021…”