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A strictly anaerobic, spore-forming bacterium (3.0-5.0 x 04-0.8 pm), designated strain SR3l (T= type strain), which stained Gram-positive and possessed a Grampositive type cell wall was isolated from a methanogenic pilot-scale digester fed with olive mill wastewater (Sfax, Tunisia). It utilized a number of carbohydrates (glucose, fructose, sorbose, galactose, myo-inositol, sucrose, lactose, cellobiose), organic compounds (lactate, betaine, sarcosine, dimethylglycine, methanethiol, dimethylsulfide), alcohol (methanol) and all methoxylated aromatic compounds only in the presence of yeast extract (0.1 %). The end products from carbohydrate fermentation were H , , CO, , formate, acetate and ethanol, that from lactate was methanol, those from methoxylated aromatics were acetate and butyrate, and that from betaine, sarcosine, dimethylglycine, methanethiol and dimethylsulfide was only acetate. Strain SR3T was non-motile, had a G+C content of 44 mol% and grew optimally at 37 "C and pH 7.4 on a glucose-containing medium. Phylogenetically, the closest relatives of strain SR3T were the nonmet hox y I at ed aroma t ic-deg rad in g Clos tridium xylanolyticum , Clostridium aerotolerans, Clostridium sphenoides and Clostridium celerecrescens (mean similarity of 98%). On the basis of the phenotypic, genotypic and phylogenetic characteristics of the isolate, it is proposed to designate strain SR3l as Clostridium methoxybenzovorans sp. nov. The type strain is SR3T (= DSM 121823.Keywords: Clostridium methoxybenzovorans, o-demethylation, aromatic compounds, syringic acid, methoxyl group INTRODUCTIONA heterogeneous collection of methoxylated aromatic compounds forms the major components of plant cell walls and these compounds are widespread in nature.A number of these compounds are degraded by some anaerobic, acetogenic and homoacetogenic bacteria. Sporobacter termitidis (Grech-Mora et al., 1997) and Holophaga foetida (Liesack et al., 1994) are two such homoacetogens that cleave the rings of the methoxylated aromatic compounds syringate, sinapate, 3,4,5-The GenBank accession number for the 165 rDNA sequence of strain SR3T is AF067965.trimethoxycinnamate and 3,4,5-trimethoxybenzoate to produce acetate, methanethiol and dimethylsulfide. In these cases, o-demethylation is the key reaction preceding aromatic ring cleavage. However, other acetogenic bacteria have been described that demethylate aromatic compounds to their corresponding hydroxylated derivatives and gain energy by the conversion of the o-methyl group to acetic acid, but are unable to cleave the aromatic ring. Examples of such cases include Sporomusa termitida (Breznak et al., 19SS), Spor-omusa malonica (Dehning et al., 1989), Sporomusa silvacetica (Kuhner et al., 1997), Moorella thermoautotrophica (formerly Clostridium thermoautotrophicum) (Collins et al., 1994; Wiegel et al., 19Sl) T. Mechichi and others aceticum) (Collins et al., 1994; Fontaine et al., 1942) and Acetobacterium woodii (Balch et al., 1977). METHODSSources of samples and organisms. Liquid samples from a...

Partial sequences of 23S rRNA gene PCR products from 23 strains of 6 pathogenic Leptospira genospecies and from 8 strains of the saprophytic Leptospira biflexa were determined. Sequence analyses enabled Leptospira genus-specific amplification primers and pathogen-specific fluorogenic adjacent hybridization probes to be designed and synthesized. A PCR protocol was developed in which changes in fluorescence emission resulting from specific annealing of fluorogenic adjacent hybridization probes to the target DNA were continuously monitored. Nine strains of the pathogenic Leptospira genospecies could be differentiated from Leptonema illini, Escherichia coli, and eight strains of Leptospira biflexa. The PCR method was rapid, requiring 18 min for the completion of 45 cycles. It was also simple and flexible, as DNA templates prepared by four different methods, including the simple boiling method, could be used without adverse effects. Two hundred copies of target, equivalent to 100 cells, could be detected.

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