Jürgen H. Thiele

Formate and ethanol are the major products of glycerol fermentation produced by a Klebsiella planticola strain isolated from red deer

The rumen contents of red deer (Cervus elaphus) were used to isolate bacteria capable of fermenting glycerol. The biochemistry, physiology, morphology and phylogeny of one isolate were studied in detail. The isolate (DR3) was tentatively identified as a strain of the species Klebsiella planticola as based on phenotypic characterization. The data obtained from 16S rRNA sequence analysis showed that the deer rumen isolate DR3 was 99·7% similar to the type strain of Kl. planticola (DSM 3069T), thus confirming the results of the phenotypic characterization. During active cell growth, it was established that glycerol dissimilation by Kl. planticola DR3 led to the production of formate and ethanol at equimolar levels of 32 mmol l−1 and 30 mmol l−1, respectively. As a result of the data obtained, a closed carbon balance was constructed for Kl. planticola DR3. This finding represented the first report of the complete end‐product profile for glycerol dissimilation by a strain of Kl. planticola isolated from cervine rumen contents.

Isolation and characterisation of obligately anaerobic, lipolytic bacteria from the rumen of red deer

Two Gram-positive, obligately anaerobic, lipolytic bacteria, isolates LIP4 and LIP5, were obtained from the rumen contents of juvenile red deer. These mesophilic bacterial strains were capable of hydrolysing the neutral lipids, tallow, tripalmitin and oliver oil, into their constituent free long-chain fatty acid and glycerol moieties. The latter compound was dissimilated by both isolates, with isolate LIP4 producing propionate as the predominant product, while isolate LIP5 produced acetate, ethanol and succinate. The lactate-utilising isolate LIP4 grew on a limited range of saccharide substrates including glucose, fructose and ribose, and exhibited an unusual cell wall structure and morphology. The isolate LIP5 grew upon a wider range of saccharides, but was unable to use lactate as a substrate. Based upon phenotypic and 16S rRNA gene sequence analyses, isolate LIP4 clusters with species in the genus Propionibacterium, while isolate LIP5 is a member of clostridial cluster XIVa.

Qualitative Rhodamine B assay which uses tallow as a substrate for lipolytic obligately anaerobic bacteria

Abstract A variety of aerobic, lipolytic bacteria and fungi have been studied in detail, due in part to increased industrial demand for lipases. However, few studies have been undertaken on obligately anaerobic lipolytic bacteria as potential sources of lipases. Using a modified version of an agar plate Rhodamine B fluorescence assay, we were able to qualitatively screen obligately anaerobic bacteria from an anaerobic digestor and the rumen of red deer for lipolytic activity towards the neutral lipids olive oil and tallow. Quantitative analysis of tallow from the agar plates inoculated with obligately anaerobic bacteria confirmed that lipolysis had occurred, and in one case, indicated that up to 82.2% of the tallow had been hydrolysed by the lipolytic activity of the bacterial strain. The qualitative tallow–Rhodamine B assay system therefore has potential as a method of screening for obligately anaerobic lipolytic bacteria using solid neutral lipids.

Isolation and characterization of two glycerol-fermenting clostridial strains from a pilot scale anaerobic digester treating high lipid-content slaughterhouse waste.

Two obligately anaerobic bacterial strains were isolated from the contents of a pilot scale, anaerobic digester treating slaughterhouse waste with a high protein and lipid content. The isolates, LIP1 and MW8, were characterized as spore‐forming, Gram‐positive rods, capable of fermenting glycerol. Isolate LIP1 was also observed to be lipolytic and was able to hydrolyse tallow and olive oil. Both isolates grew optimally at 37 °C and formed either acetate and formate (LIP1), or acetate and butyrate (MW8), as major glycerol fermentation products. Both isolates produced ethanol as the major reduced fermentation end‐product. Neither MW8 nor LIP1 had growth and metabolism inhibited by the addition of stearic acid at concentrations normally considered bactericidal. Analysis of the 16S rRNA gene sequences, in conjunction with the phenotypic data, confirmed that the isolates are members of the genus Clostridium (sensu lato), clustering with species of clostridial clusters I (MW8) and XIVa (LIP1).

High-performance liquid chromatographic analysis of free long chain fatty acids produced during lipolysis by anaerobic digestor sludge

Abstract The development of sample extraction techniques in conjunction with application of a modified version of an established HPLC technique allowed the rapid estimation of concentrations of long chain fatty acids (LCFA) produced by the lipolytic activity of bacteria in anaerobic digestor sludges or experiments using the sludges as inocula. It was established that free long chain fatty acids will preferentially partition into certain phases in the sludge or experimental cultures. These data, and application of the developed techniques for monitoring of LCFA, may lead to the avoidance of toxicity and failure of full-scale anaerobic digestors treating high lipid content waste in the future.

Modification of an algal culture medium for sustained growth of a saxitoxin-producing isolate of Alexandrium minutum

In order to study saxitoxin (STX) production bymicro-algae in the laboratory, a defined algal culture medium which supports optimum growth over a longtime-period is a requirement. In the development of such a medium, a number of modifications were made to a standard algal culture medium (GP) and growth of a STX-producing isolate of Alexandrium minutum in the different formulations was assessed by measuring maximum cell densities and mean generation times (MGT). All experiments were carried out under controlled conditions in an aerobic atmosphere with increased CO2. Whilst maximum cell densities in the different modifications were similar, the MGT was significantly shortened by the addition of Tris buffer and the trace metals strontium, selenium and molybdenum. Replacement of natural with artificial seawater and removal of soil extract did not adversely affect algal growth. Five of the six media formulations supported the growth of A. minutumover a 9-month period.

Recalcitrance of DDE (2,2‐BIS‐(4‐chlorophenyl)‐1,1 dichloroethylene) and DPE (1,1‐diphenylethylene) to cometabolic aerobic biodegradation

Bacterial degradation of the persistent DDT [1,1,1‐trichloro‐2,2‐bis(4‐chloro‐pheny‐l)ethane] metabolite DDE has not previously been reported. Bacteria from DDT, PCP and PAH contaminated soils and sediments were extracted and evaluated for their ability to degrade DDE and its dehalogenated derivative DPE. Aerobic aromatic hydrocarbon degrading bacteria were enriched using chemostat techniques under both carbon limited and nitrogen limited selection conditions. DDE, ethylbenzene (EB), DPE and dipyridyl ketone oxime (DPKO) were added as structurally related potentially biodegradable carbon and nitrogen sources. No significant degradation of DDE and DPE was observed under these conditions. Pure cultures isolated from the enrichments on nonselective growth media were able to utilise EB and DPKO as carbon and nitrogen sources, respectively but were unable to degrade DDE or DPE alone or in the presence of EB and DPKO. The absence of DDE degradation capacity in the microbial community was confirmed by adding 14C...