- Volume 49, Issue 4, 1999

Two aerobic, Gram-positive bacteria, strains HKI 0122T and HKI 0132, were isolated from a cave. Cells are not acid-fast, non-motile, non-spore-forming and exhibit a rod-coccus growth cycle. The cell wall peptidoglycan contains lysine in position 3 of the peptide subunit and an interpeptide bridge of l-Lys ← l-Glu. The major menaquinone is MK-8(H4), 13-methyl and 12-methyl tetradecanoic acids are the predominating fatty acids. The polar lipids consist of phosphatidylinositol, diphosphatidylglycerol and three unknown phospholipids. Mycolic acids are absent. The DNA base composition is 71 mol% G+C. Phylogenetic analysis revealed that strain HKI 0122T forms a novel taxon among the families and unassigned genera of the suborder Micrococcineae, within the order Actinomycetales. On the basis of the genotypic, chemotaxonomic, morphological and physiological characteristics of these two isolates it is proposed to assign strains HKI 0122T and HKI 0132 to a new genus and species for which the name Beutenbergia cavernae gen. nov., sp. nov. is proposed. The type strain is HKI 0122T (= DSM 12333T).

A strictly chemo-organotrophic, anaerobic bacterium was isolated from an olive mill wastewater treatment digester on syringate and designated strain SR1T. The cells were slightly curved rods, stained Gram-positive and possessed terminal spores. Strain SR1T utilized crotonate, methanol and a wide range of aromatic compounds including 3,4,5-trimethoxybenzoate (TMB), 3,4,5-trimethoxycinnamate (TMC), syringate, 3,4,5-trimethoxyphenylacetate (TMPA), 3,4,5-trimethoxyphenylpropionate (TMPP), ferulate, sinapate, vanillate, 3,4-dimethoxybenzoate, 2,3-dimethoxybenzoate, gallate, 2,4,6-trihydroxybenzoate (THB), pyrogallol, phloroglucinol and quercetin as carbon and energy sources. Acetate and butyrate were produced from aromatic compounds, methanol and crotonate whereas methanethiol (MT) was produced from methoxylated aromatic compounds and methanol. Strain SR1T had a G+C content of 38 mol% and grew optimally between 37 and 40 °C at pH 7·2 on a crotonate-containing medium. Phylogenetically, strain SR1T was a member of cluster XIVa of the Clostridiales group and shared a sequence similarity of 90% with Clostridium aminovalericum and Eubacterium fissicatena. Consequently, its precise neighbourliness to any one of them depended on the selection of strains of the cluster. On the basis of the phylogenetic and phenotypic evidence presented in this paper, the designation of strain SR1T as Sporobacterium olearium gen. nov., sp. nov. is proposed. The type strain is SR1T (= DSM 12504T).

Two actinomycete strains, DF-28 and DF-32T, were isolated from soil samples collected in a deciduous dipterocarp forest in Thailand. They produced longitudinally paired spores on the tips of short sporophores alternately branched from aerial hyphae, and the chemotaxonomic properties of the isolates were the same as those of members of the family Streptosporangiaceae. These phenotypic properties, together with the results of a phylogenetic analysis based on 16S rRNA gene sequences, indicated that these isolates should be assigned to the genus Microbispora. The two isolates showed more than 93% DNA relatedness to each other, but their relatedness to any previously described species of the genus Microbispora was only 45% or less. They were distinguishable from previously described Microbispora spp. by a combination of physiological and biochemical properties. Therefore, a new species is proposed for these strains, under the name Microbispora corallina sp. nov. The type strain is strain DF-32T (= JCM 10267T).

A strain was isolated from soil that formed short chains of aerial mycelium dividing into motile spores on an agar medium, but not substrate mycelium. The organism had menaquinone 9(H8), LL-diaminopimelic acid in the cell wall amino acids and a DNA G+C content of 71 mol%. On the basis of the morphological and chemotaxonomic characteristics and phylogenetic analysis, this strain was classified into the genus Sporichthya. DNA-DNA hybridization revealed that the strain differed from the known species, Sporichthya polymorpha. Therefore, a new species, Sporichthya brevicatena sp. nov., is proposed. The type strain is S. brevicatena YU720-21T (= IFO 16195T).

A rubber-degrading bacterium (strain Kd21) was isolated from fouling tyre water inside a deteriorated automobile tyre. The strain was aerobic, Grampositive, produced elementary branching hyphae which fragmented into rod/coccus-like elements and showed chemotaxonomic markers which were consistent with the classification of Gordonia, i.e. meso-diaminopimelic acid, N-glycolyl muramic acid, arabinose and galactose as diagnostic sugars, a fatty acid pattern composed of unbranched saturated and monounsaturated fatty acids with a considerable amount of tuberculostearic acid, and mycolic acids comprising 58-66 carbon atoms with two principal mycolic acids C60 and C62 counting for over 60%. Results of 16S rDNA analyses as well as chemotaxonomic results, led to the conclusion that Gordonia sp. strain Kd2T (= DSM 44302T) represents a new species within the genus Gordonia for which the name Gordonia polyisoprenivorans is proposed.

Strain GrollT, isolated from fresh water, is a mesophilic, spore-forming, sulfatereducing bacterium that uses a large variety of substrates as electron donors ranging from simple organic compounds to long-chain fatty acids and several aromatic compounds. Sulfate, thiosulfate and sulfite are used as electron acceptors. Homoacetogenic growth occurs under sulfate-free conditions. Substrate oxidation is usually complete, leading to CO2, but acetate or other fatty acids can accumulate at high substrate concentrations. The G+C content of the DNA is 54·8 mol%. Strain GrollT was found to be phenotypically and phylogenetically different from known members of the genus Desulfotomaculum. 16S rRNA gene sequence analyses show that this organism falls within the radiation of the genus Desulfotomaculum cluster and has <96% sequence similarity to previously described species. The name Desulfotomaculum gibsoniae sp. nov. is proposed for this strain; the type strain is GrollT (= DSM 7213T).

A new, slow-growing, scotochromogenic mycobacterium was isolated from a lymph node of an immunocompromised child and subsequently from tap water and from a respiratory specimen of a patient with chronic fibrosis. Alcohol-acid-fastness, lipid patterns and the G+C content clearly support the placement of this organism in the genus Mycobacterium. The isolates grew very slowly at temperatures ranging from 25 to 32 °C and showed activities of nitrate reductase, catalase, urease, arylsulfatase and Tween 80 hydrolysis. The organism was susceptible to all antimycobacterial drugs tested. The 16S rDNA sequence was unique and phylogenetic analysis placed the organism close to fast-growing species such as Mycobacterium farcinogenes, Mycobacterium komossense and Mycobacterium aichiense. These data support the conclusion that the isolates represent a new mycobacterial species, for which the name Mycobacterium tusciae sp. nov. is proposed. The type strain is strain Fl-25796T; a culture of this strain has been deposited in the DSMZ as strain DSM 44338T.

The taxonomic position of two actinomycetes isolated from soil was established using a polyphasic approach. The organisms, designated 213ET and 213F, were found to have chemical and morphological properties consistent with their assignment to the genus Gordonia. Nearly complete sequences of the 16S rDNA genes of the two strains were determined following the isolation and direct sequencing of the amplified genes. The tested strains were found to have identical 16S rDNA sequences and formed a phylogenetic line within the evolutionary radiation occupied by the genus Gordonia that was most closely related to Gordonia rubropertincta DSM 43197T. However, DNA-DNA relatedness data showed that strain 213ET and Gordonia rubropertincta DSM 43197T belonged to distinct genomic species. Strains 213ET and 213F also shared an identical phenotypic profile which distinguished them from representatives of validly described Gordonia species. The combined genotypic and phenotypic data show that strains 213ET and 213F merit recognition as a new species of Gordonia. The name proposed for the new species is Gordonia desulfuricans, for which the type strain is 213ET (= NCIMB 40816T).

The taxonomic position of the genus Planopolyspora comprising a single species, Planopolyspora crispa, was reviewed. This genus was originally characterized by formation of long, curly and sometimes branching sporangia containing numerous zoospores arranged in a single row and by the presence of meso-diaminopimelic acid and madurose (3-O-methyl-d-galactose) in wholecell hydrolysates. However, our chemotaxonomic analyses of the type strain of P. crispa did not agree with the original description. The peptidoglycan contained l-lysine but not meso-diaminopimelic acid, and the whole-cell hydrolysate contained xylose as the characteristic sugar but not madurose. These characteristics and other chemotaxonomic profiles (e.g. menaquinone, phospholipid and cellular fatty acid compositions) of the genus Planopolyspora coincided with those of the genus Catenuloplanes. These two genera also had very similar morphological characteristics, but in the original description of the genus Catenuloplanes the presence of sporangia was not referred to. This difference is considered to originate from a divergence of views owing to the ambiguity of the definition of the term ‘sporangium’ in actinomycete morphology. Phylogenetic analysis based on 16S rDNA sequences also supported the proposal that the genera Planopolyspora and Catenuloplanes should be combined into one genus. Levels of DNA relatedness among the type strains of P. crispa and six Catenuloplanes species and their cultural, physiological and biochemical characteristics indicated that P. crispa should be classified as an independent species of the genus Catenuloplanes, which has priority over the genus Planopolyspora. Therefore, it is proposed that Planopolyspora crispa be transferred to the genus Catenuloplanes as Catenuloplanes crispus comb. nov.

Chemotaxonomic, electron microscopic and 16S rRNA gene sequence analyses of the three described species of the genus Anaerovibrio demonstrated only remote similarities to each other. The 16S rRNA gene sequence similarities between Anaerovibrio lipolytica, Anaerovibrio glycerini and Anaerovibrio burkinabensis and the derived phylogenetic relationships of the three specie studied fell below genus level. All three species clustered within the Sporomusa-Pectinatus-Selenomonas phyletic group. Each species showed a distinct phospholipid pattern and whole-cell fatty acid distribution. Several isoprenologues of the lipoquinone ‘lipid F’ were found to differ in their quantitative distribution in the Anaerovibrio species. On the basis of these results, the new genera Anaerosinus gen. nov. and Anaeroarcus gen. nov. are proposed. The type species of Anaerosinus is Anaerosinus glycerini comb. nov., and the type species of Anaeroarcus is Anaeroarcus burkinensis [corrig.] comb. nov. The genus Anaerovibrio is consequently restricted to a single species, namely Anaerovibrio lipolyticus [corrig.].

Four strains of a previously undescribed Actinomyces-like bacterium were isolated from canine and feline clinical specimens. Phenotypic studies indicated the strains were members of the genus Actinomyces, and most closely resembled Actinomyces viscosus serotype I and Actinomyces slackii. Comparative 16S rRNA gene sequencing studies demonstrated the unknown bacterium constitutes a new subline within a group of Actinomyces species, which includes Actinomyces bovis, the type species of the genus. Based on phylogenetic and phenotypic evidence it is proposed that the unknown bacterium be classified as Actinomyces bowdenii sp. nov. The type strain of Actinomyces bowdenii is CCUG 37421T.

Strain K1T, isolated from the gastrointestinal tract of Atlantic salmon (Salmo salar), has the capacity to inhibit the growth of the fish pathogens Vibrio anguillarum and Aeromonas salmonicida. Strain K1T is a motile Gram-positive psychrophilic rod that lacks both catalase and oxidase, which does not grow on acetate containing media, but grows at pH 9 and in TSB with up to 6 % sodin chloride content. Strain K1T is facultatively anaerobic and tryptone as a sole source of nutrient promotes growth. The most abundant cellular fatty acid of strain K1T is oleic acid (18:1cis9). Based on 16S rDNA sequence comparisons, it is suggested that strain K1T is phylogenetically closely related to C. alterfunditum. However, the unique phenotypic attributes of strain K1T suggest that it represents a new species. The name Carnobacterium inhibens i proposed, for which the type strain is K1T (= CCUG 31728T).

Seven strains of three new yeast species were isolated from soil, flowers and leaves in the Nansei Islands, Japan. These isolates most closely resembled Kluyveromyces phaffii in physiological characteristics and nuclear DNA base composition (30–32 mol% G+C), but on the basis of DNA-DNA hybridization and electrophoretic karyotyping they were categorized into three new species different from K. phaffii. Phylogenetic analysis using 18S rRNA gene sequence showed that the three new species and K. phaffii were highly related to one another and phylogenetically separate from the members of other species. On the basis of phylogeny and physiological characters, it is proposed that the three new species represent novel taxa and should be designated Tetrapisispora iriomotensis gen. nov., sp. nov. (type strain IFO 10929T), Tetrapisispora nanseiensis gen. nov., sp. nov. (type strain IFO 10899T) and Tetrapisispora arboricola gen. nov., sp. nov. (type strain IFO 10925T), while Kluyveromyces phaffii becomes Tetrapisispora phaffii comb. nov.

The 16S rDNA sequences of the recently described Aeromonas encheleia and Aeromonas popoffii, were determined and compared with data from all known Aeromonas sp. Diagnostic 16S rDNA regions were also sequenced for some strains previously considered as an extension of A. encheleia and a strain of Aeromonas Group 501 (formerly Enteric Group 501). Results indicated that A. encheleia and A. popoffii are phylogenetically separated species as originally described. A. conclusion about HG11 taxonomic status is not recommended until previous discrepancies are clarified by further DNA-DNA hybridization and sequencing studies.

The 16S rRNA sequences of the Korean Borrelia strains 5MT and 9MT, isolated from lxodes nipponensis, showed identities of 99.0-99.1% to that of B. afzelii. The strains were tentatively classified as belonging to the B. afzelii-related group. In this study, Korean isolates, including these strains, were characterized further and compared with recently described new species. These strains generated a RFLP pattern that has not been found previously in RFLP analysis of the 5S-23S rRNA intergenic spacer and the flagellin gene. When phylogenetic trees were constructed, based on the 5S-23S rRNA intergenic spacer, flagellin gene and 16S rRNA sequences, these Korean isolates formed a cluster with the Borrelia strain Am501 isolated from Ixodes columnae in Japan and Borrelia valaisiana strains VS116T and UK isolated from Ixodes ricinus in Europe and were distinguishable from the other species. However, these three groups of strains were divergent from each other in the molecular masses of the putative outer surface protein A (OspA) and in the sequences of the ospA gene. These findings suggest that these Korean isolates and one Japanese isolate are members of B. valaisiana and that OspA of this species is divergent, as is that of Borrelia garinii. This led to the speculation that B. valaisiana strains are adapted to the vector ticks found in each locality.

DNA sequences covering 36% of the mle gene that encodes the malolactic enzyme were determined for 13 strains of lactic acid bacteria, representing Pediococcus, Leuconostoc, Lactobacillus and Oenococcus genera. The sequences were aligned with the corresponding region of mleS in Lactococcus lactis. The phylogenetic distance matrix tree of all mle sequences was compared with the 16S rRNA phylogenetic tree. The analysis showed that the mle fragment evolved more rapidly than the 16S gene and differently. Pediococcus and Lactobacillus species were intermixed in the 16S rRNA tree whereas they were separated in the mle tree. Leuconostoc mesenteroides and Oenococcus oeni were distinct from other species in the 16S rRNA tree, whereas they were intermixed with Lactobacillus species and Lactococcus lactis in the mle tree. The amino acid sequences deduced from partial mle genes were aligned with 22 malic enzyme sequences and the corresponding phylogenetic tree was constructed. Malic and malolactic enzymes were distinct at the phylogenetic level, except for malic enzymes of yeast and Escherichia coli which were nearer the malolactic enzymes than the other malic enzymes. The analysis of conserved sites showed several interesting amino acids specific to either malic enzyme or malolactic enzyme.

Nodule isolates from 11 species of wild legumes in north-western China were characterized by numerical taxonomy, PCR-based 16S rRNA gene RFLP and sequence analyses, DNA-DNA hybridization, restriction patterns of nodDAB and nifH genes, and symbiotic properties. Based on the results of numerical taxonomy, most of the 35 new isolates were grouped into five clusters (clusters 7, 9, 12, 14 and 15). Clusters 7 and 12 were identified as Mesorhizobium amorphae and Agrobacterium tumefaciens, respectively, based on their high DNA homologies with the reference strains for these species, their 16S rRNA gene analysis and their phenotypic features. Results of 16S rDNA PCR-RFLP analysis showed that cluster 9 belonged to Rhizobium. Clusters 14 and 15 were identified as Mesorhizobium based on their moderately slow-growing, acid-producing characters and the high similarity of their 16S rDNA PCR-RFLP patterns to those of Mesorhizobium species. These two clusters were genomic species distinct from all described species based on analysis of DNA relatedness within this genus. The isolates in cluster 12 (Agrobacterium tumefaciens) failed to nodulate their original host and other selected hosts and they did not hybridize to nif or nod gene probes. The possibility of opportunistic nodulation of these isolates is discussed. Identical restriction patterns were obtained in the nif or nod gene hybridization studies from the three isolates within cluster 15, which were isolated from the same host species. The isolates from different host plants in each of clusters 9 and 14 produced different nodDAB RFLP patterns, but similar nifH RFLP patterns appeared (one band for each isolate). Different patterns were observed among different clusters from both the nod and nif gene hybridization studies. Cross-nodulation was recorded among the isolates and the host plants in the same cluster and promiscuous properties were found among some of the hosts tested.