- Volume 55, Issue 5, 2005

A novel strictly aerobic, heterotrophic, pink-pigmented, non-motile, Gram-negative, oxidase-, catalase-, β-galactosidase- and alkaline phosphatase-positive marine bacterium, designated strain KMM 6058T, was isolated from the sea urchin Strongylocentrotus intermedius and studied using a polyphasic taxonomic approach. The G+C content of the DNA of the isolate was 41·3 mol%. The predominant fatty acids were i15 : 1, i15 : 0, a15 : 0 and i17 : 0 3-OH. Phylogenetic analysis based on 16S rRNA gene sequences revealed that strain KMM 6058T formed a monophyletic clade with Roseivirga ehrenbergii, with 99 % similarity. On the basis of phenotypic, chemotaxonomic, genotypic and phylogenetic characteristics, the novel bacterium should be assigned to the genus Roseivirga as Roseivirga echinicomitans sp. nov. The type strain is KMM 6058T (=KCTC 12370T=LMG 22587T).

Six bacterial strains isolated from the human oral cavity, PPPA16, PPPA20T, PPPA24, PPPA31, EPPA6 and EPPA7, were characterized by determining phenotypic and biochemical features, cellular fatty acid profiles, menaquinone profiles and phylogenetic position based on 16S rRNA gene sequence analysis. 16S rRNA gene sequence analysis showed that the isolates represented the same species of the genus Prevotella. The strains were related to Prevotella dentalis with about 89 % similarity. In addition, the isolates were related to Prevotella sp. oral clone IDR-CEC-0032, which is a representative of the numerically dominant cluster VI in carious dentine lesions [ Nadkarni et al. (2004). J Clin Microbiol 42, 5238–5244 ], with about 99 % similarity. The strains were obligately anaerobic, non-pigmenting, non-spore-forming, non-motile, Gram-negative rods. The isolates could be differentiated from other Prevotella species by d-mannitol, d-melezitose, d-sorbitol and d-trehalose fermentation in API 20A tests. The cellular fatty acid composition of strains PPPA16, PPPA20T, PPPA24, PPPA31, EPPA6 and EPPA7 was significantly different from that of other Prevotella species. Compared with other Prevotella species, only these six strains contained dimethyl acetals. The major menaquinones of the clinical isolates were MK-12 and MK-13, whereas the major menaquinones of other Prevotella species were MK-10 and MK-11. On the basis of these data, a novel Prevotella species, Prevotella multisaccharivorax sp. nov., is proposed, with PPPA20T (=JCM 12954T=DSM 17128T) as the type strain.

Three Gram-negative, yellow-pigmented, rod-shaped bacterial strains, SMK-12T, SMK-36 and SMK-45, were isolated from a tidal flat sediment of the Yellow Sea in Korea, and their taxonomic positions were investigated by a polyphasic approach. The three strains grew optimally at 25–30 °C and in the presence of 2–3 % (w/v) NaCl. They contained MK-6 as the predominant menaquinone. The major cellular fatty acids were iso-C15 : 0, iso-C17 : 0 3-OH, iso-C15 : 1, anteiso-C15 : 0, iso-C15 : 0 3-OH and C16 : 1 ω7c and/or iso-C15 : 0 2-OH. The DNA G+C contents of the three strains were 34·7–34·9 mol%. The phylogenetic tree based on 16S rRNA gene sequences revealed that the three strains form one distinct evolutionary lineage supported by a bootstrap value of 100 % within the family Flavobacteriaceae. The three strains exhibited 16S rRNA gene sequence similarity levels of 93·8–94·9 % to the nearest phylogenetic neighbours, the genera Algibacter, Bizionia and Formosa. On the basis of differences in phenotypic characteristics and phylogenetic distinctiveness, strains SMK-12T, SMK-36 and SMK-45 were classified in a novel genus and species, for which the name Gaetbulibacter saemankumensis gen. nov., sp. nov. is proposed. The type strain for the novel species is SMK-12T (=KCTC 12379T=DSM 17032T).

The obligately intracellular coccoid bacterium UWE25, a symbiont of Acanthamoeba spp., was previously identified as being related to chlamydiae based upon the presence of a chlamydia-like developmental cycle and its 16S rRNA gene sequence. Analysis of its complete genome sequence demonstrated that UWE25 shows many characteristic features of chlamydiae, including dependency on host-derived metabolites, composition of the cell envelope and the ability to thrive as an energy parasite within the cells of its eukaryotic host. Phylogenetic analysis of 44 ribosomal proteins further confirmed the affiliation of UWE25 to the ‘Chlamydiae’. Within this phylum, UWE25 could be assigned to the family Parachlamydiaceae based on comparative analyses of the 16S rRNA, 23S rRNA and endoribonuclease P RNA genes. The distinct dissimilarities from its closest relative, Parachlamydia acanthamoebae Bn9 T (7·1, 9·7 and 28·8 %, respectively), observed in this analysis justify its classification in a new genus. Therefore, the name ‘Candidatus Protochlamydia amoebophila’ is proposed for the designation of the Acanthamoeba sp. symbiont UWE25 (=ATCC PRA-7).

A yellow-pigmented bacterium designated strain GUM-KajiT was isolated from a lactic acid beverage. The strain had Gram-negative, non-motile, rod-shaped cells. It was strictly aerobic and chemo-organotrophic and grew at 5–30 °C and at pH 5–8. The major components of the non-polar and 3-hydroxy fatty acids were C15 : 0 iso and 3-OH-C17 : 0 iso, respectively. Menaquinone MK-6 was detected as the sole quinone. 16S rRNA gene sequence comparisons revealed that strain GUM-KajiT is affiliated to the genus Chryseobacterium, with Chryseobacterium joostei as its phylogenetic neighbour, but there were low levels of similarity (<96 %) to any established species of the genus. The G+C content of the genomic DNA was 36·6 mol%. The novel bacterium differed from any known species of Chryseobacterium in terms of a number of phenotypic properties. Thus, the name Chryseobacterium shigense sp. nov. is proposed for this novel bacterium. The type strain is strain GUM-KajiT (=BAMY 1001T=NCIMB 14047T=DSM 17126T).

Two Gram-negative, non-spore-forming, slightly halophilic gliding bacterial strains, DSW-8T and DSW-9, were isolated from sea water off a Korean island, Dokdo, of the East Sea, Korea, and their taxonomic position was investigated by a polyphasic study. The two strains grew optimally at 30 °C and in the presence of 2–3 % (w/v) NaCl. Strains DSW-8T and DSW-9 were characterized chemotaxonomically as containing MK-6 as the predominant menaquinone and iso-C17 : 0 3-OH, iso-C15 : 0 and iso-C15 : 1 as the major fatty acids. Major polar lipids were phosphatidylethanolamine, two unidentified phospholipids, an unidentified glycolipid and an amino group-containing lipid that was ninhydrin-positive. Their DNA G+C contents were 36·1 and 35·9 mol%. Phylogenetic analyses based on 16S rRNA gene sequences showed that strains DSW-8T and DSW-9 fell within the genus Maribacter of the family Flavobacteriaceae. Strains DSW-8T and DSW-9 exhibited no difference in their 16S rRNA gene sequences and possessed a mean DNA–DNA relatedness level of 89 %. Strains DSW-8T and DSW-9 exhibited 16S rRNA gene sequence similarity levels of 96·9–98·0 % to the type strains of the four recognized Maribacter species, but their low level of DNA–DNA relatedness with these species demonstrated that they constitute a distinct Maribacter species. On the basis of phenotypic and phylogenetic data and genetic distinctiveness, strains DSW-8T (=KCTC 12393T=DSM 17201T) and DSW-9 were classified in the genus Maribacter as members of a novel species, for which the name Maribacter dokdonensis sp. nov. is proposed.

Nine strains of Gram-negative, anaerobic rod were isolated from human faeces. Based on phylogenetic analysis and specific phenotypic characteristics, these strains were included within the Bacteroides cluster and were divided into two clusters. Strains from the two clusters showed 16S rRNA gene sequence similarities of 90·4 and 92·7 % to the nearest recognized species, Bacteroides vulgatus. The strains also formed two clusters exhibiting a 16S rRNA gene sequence divergence of approximately 6 %. DNA–DNA hybridization studies confirmed that the two novel strain clusters were distinct from each other. Based on the phenotypic and phylogenetic findings, two novel species, Bacteroides plebeius sp. nov. and Bacteroides coprocola sp. nov., are proposed, each representing one of the two strain clusters. The DNA G+C content of the type strains were 43·9 mol% for B. plebeius (M12T=JCM 12973T=DSM 17135T) and 42·4 mol% for B. coprocola (M16T=JCM 12979T=DSM 17136T).

Yellow-pigmented, Gram-negative organisms isolated from raw chicken were investigated by means of a polyphasic taxonomic approach and were shown to represent a novel species in the genus Chryseobacterium, for which the name Chryseobacterium vrystaatense sp. nov. is proposed. Its nearest phylogenetic neighbours were Chryseobacterium joostei, Chryseobacterium indologenes and Chryseobacterium gleum, which showed 16S rRNA gene sequence similarity levels of 96·9, 97·1 and 96·1 %, respectively. Levels of DNA–DNA hybridization between strains of C. vrystaatense and Chryseobacterium reference species were below 46 %. Strain LMG 22846T (=CCUG 50970T) was chosen as the type strain and has a DNA G+C content of 37·1 mol%.

A novel, strictly aerobic, Gram-negative, yellow–orange-pigmented, motile, catalase-positive, oxidase-negative bacterium, RW262T, was isolated from water of the River Taff, Cardiff, UK, during January 2000. Phylogenetic analysis of the 16S rRNA gene indicated that strain RW262T was a member of the family Cryomorphaceae within the phylum ‘Bacteroidetes’. The DNA G+C content of strain RW262T was 37·2 mol%. The predominant fatty acid was the branched-chain saturated fatty acid i15 : 0 (44·2 %). On the basis of polyphasic analysis of phenotypic, chemotaxonomic, genotypic and phylogenetic characteristics, it is proposed that this freshwater bacterium represents a novel genus and species within the family Cryomorphaceae, Fluviicola taffensis gen. nov., sp. nov. The type strain is RW262T (=NCIMB 13979T=DSM 16823T).

Twelve chemolithotrophic strains were isolated from temperate orchard soil on reduced sulfur compounds as energy and electron sources and characterized on the basis of their physiological properties and ability to oxidize various reduced sulfur compounds. The new isolates could oxidize tetrathionate as well as thiosulfate, and oxidation of the latter involved conversion of thiosulfate to tetrathionate followed by its accumulation and eventual oxidation to sulfate, manifested in the production of acid. The mesophilic, neutrophilic, Gram-negative and coccoid bacteria had a respiratory metabolism. Physiologically and biochemically, all the strains were more or less similar, differing only in their growth rates and ability to utilize a few carbon compounds as single heterotrophic substrates. 16S rRNA gene sequence analysis was performed with five representative strains, which revealed a high degree of similarity (⩾99 %) among them and placed the cluster in the ‘Betaproteobacteria’. The strains showed low levels (93·5–95·3 %) of 16S rRNA gene sequence similarity to Pigmentiphaga kullae, Achromobacter xylosoxidans, Pelistega europaea and species belonging to the genera Alcaligenes, Taylorella and Bordetella. The taxonomic coherence of the new isolates was confirmed by DNA–DNA hybridization. On the basis of their uniformly low 16S rRNA gene sequence similarities to species of all the closest genera, unique fatty acid profile, distinct G+C content (54–55·2 mol%) and phenotypic characteristics that include efficient chemolithotrophic utilization of tetrathionate, the organisms were classified in a new genus, Tetrathiobacter gen. nov. In the absence of any significant discriminatory phenotypic or genotypic characteristics, all the new isolates are considered to constitute a single species, for which the name Tetrathiobacter kashmirensis sp. nov. (type strain WT001T=LMG 22695T=MTCC 7002T) is proposed.

A novel genus and species are proposed for two strains of methanotrophic bacteria isolated from hypersaline lakes in the Crimean Peninsula of Ukraine. Strains 10KiT and 4Kr are moderate halophiles that grow optimally at 1–1·5 M (5·8–8·7 %, w/v) NaCl and tolerate NaCl concentrations from 0·2 M up to 2·5 M (1·2–15 %). This optimum and upper limit are the highest for any methanotrophic bacterium known to date. The strains are Gram-negative, aerobic, non-pigmented, motile, coccoid to spindle-shaped bacteria that grow on methane or methanol only and utilize the ribulose monophosphate pathway for carbon assimilation. They are neutrophilic (growth occurs only in the range pH 6·5–7·5) and mesophilic (optimum growth occurs at 30 °C). On the basis of 16S rRNA gene sequence phylogeny, strains 10KiT and 4Kr represent a type I methanotroph within the ‘Gammaproteobacteria’. However, the 16S rRNA gene sequence displays <91·5 % identity to any public-domain sequence. The most closely related methanotrophic bacterium is the thermophilic strain HB. The DNA G+C content is 58·7 mol%. The major phospholipid fatty acids are 18 : 1ω7 (52–61 %), 16 : 0 (22–23 %) and 16 : 1ω7 (14–20 %). The dominance of 18 : 1 over 16 : 0 and 16 : 1 fatty acids is unique among known type I methanotrophs. The data suggest that strains 10KiT and 4Kr should be considered as belonging to a novel genus and species of type I methanotrophic bacteria, for which the name Methylohalobius crimeensis gen. nov., sp. nov. is proposed. Strain 10KiT (=DSM 16011T=ATCC BAA-967T) is the type strain.

The bacterial strains IMB-1T and CC495T, which are capable of growth on methyl chloride (CH3Cl, chloromethane) and methyl bromide (CH3Br, bromomethane), were isolated from agricultural soil in California fumigated with CH3Br, and woodland soil in Northern Ireland, respectively. Two pesticide-/herbicide-degrading bacteria, strains ER2 and C147, were isolated from agricultural soil in Canada. Strain ER2 degrades N-methyl carbamate insecticides, and strain C147 degrades triazine herbicides widely used in agriculture. On the basis of their morphological, physiological and genotypic characteristics, these four strains are considered to represent two novel species of the genus Aminobacter, for which the names Aminobacter ciceronei sp. nov. (type strain IMB-1T=ATCC 202197T=CIP 108660T=CCUG 50580T; strains ER2 and C147) and Aminobacter lissarensis sp. nov. (type strain CC495T=NCIMB 13798T=CIP 108661T=CCUG 50579T) are proposed.

A novel Ferrimonas species is described on the basis of phenotypic, chemotaxonomic and phylogenetic studies. Four halophilic organisms were isolated from marine sand and marine macroalgae samples by using high-pH marine agar 2216. An analysis of the nearly complete 16S rRNA gene sequences of these new isolates indicated that they were phylogenetically close (16S rRNA gene sequence similarity >99·5 %, gyrB gene sequence similarity >97·8 %), and were most closely related to Ferrimonas balearica (16S rRNA gene sequence similarity 97·1–97·3 %, gyrB gene sequence similarity 84·4–85·0 %). Chemotaxonomic data (major menaquinone MK7; major fatty acids C16 : 0 and C18 : 1 ω9c) supported the affiliation of the new isolates to the genus Ferrimonas. The results of physiological and biochemical tests allowed phenotypic differentiation of the isolates from F. balearica. It is therefore proposed that the new isolates represent a novel species with the name Ferrimonas marina sp. nov. and type strain A4D-4T (=MBIC06480T=DSM 16917T).

Symptoms of huanglongbing (HLB) were reported in São Paulo State (SPS), Brazil, in March 2004. In Asia, HLB is caused by ‘Candidatus Liberibacter asiaticus' and in Africa by ‘Candidatus Liberibacter africanus’. Detection of the liberibacters is based on PCR amplification of their 16S rRNA gene with specific primers. Leaves with blotchy mottle symptoms characteristic of HLB were sampled in several farms of SPS and tested for the presence of liberibacters. ‘Ca. L. asiaticus' was detected in a small number of samples but most samples gave negative PCR results. Therefore, a new HLB pathogen was suspected. Evidence for an SPS-HLB bacterium in symptomatic leaves was obtained by PCR amplification with universal primers for prokaryotic 16S rRNA gene sequences. The amplified 16S rRNA gene was cloned and sequenced. Sequence analysis and phylogeny studies showed that the 16S rRNA gene possessed the oligonucleotide signatures and the secondary loop structure characteristic of the α-Proteobacteria, including the liberibacters. The 16S rRNA gene sequence phylogenetic tree showed that the SPS-HLB bacterium clustered within the α-Proteobacteria, the liberibacters being its closest relatives. For these reasons, the SPS-HLB bacterium is considered a member of the genus ‘Ca. Liberibacter’. However, while the 16S rRNA gene sequences of ‘Ca. L. asiaticus' and ‘Ca. L. africanus' had 98·4 % similarity, the 16S rRNA gene sequence of the SPS-HLB liberibacter had only 96·0 % similarity with the 16S rRNA gene sequences of ‘Ca. L. asiaticus' or ‘Ca. L. africanus’. This lower similarity was reflected in the phylogenetic tree, where the SPS-HLB liberibacter did not cluster within the ‘Ca. L asiaticus’/‘Ca. L. africanus group’, but as a separate branch. Within the genus ‘Candidatus Liberibacter’ and for a given species, the 16S/23S intergenic region does not vary greatly. The intergenic regions of three strains of ‘Ca. L. asiaticus’, from India, the People's Republic of China and Japan, were found to have identical or almost identical sequences. In contrast, the intergenic regions of the SPS-HLB liberibacter, ‘Ca. L. asiaticus' and ‘Ca. L. africanus' had quite different sequences, with similarity between 66·0 and 79·5 %. These results confirm that the SPS-HLB liberibacter is a novel species for which the name ‘Candidatus Liberibacter americanus' is proposed. Like the African and the Asian liberibacters, the ‘American’ liberibacter is restricted to the sieve tubes of the citrus host. The liberibacter could also be detected by PCR amplification of the 16S rRNA gene in Diaphorina citri, the psyllid vector of ‘Ca. L. asiaticus’, suggesting that this psyllid is also a vector of ‘Ca. L. americanus' in SPS. ‘Ca. L. americanus' was detected in 216 of 218 symptomatic leaf samples from 47 farms in 35 municipalities, while ‘Ca. L. asiaticus' was detected in only 4 of the 218 samples, indicating that ‘Ca. L. americanus' is the major cause of HLB in SPS.

A novel moderately thermophilic methanotroph, strain MYHTT, was isolated from a hot spring in Japan. The isolate grew on methane or methanol at 37–67 °C, and optimally at 57–59 °C. It was found to be a Gram-negative aerobe, with colourless colonies of non-motile coccoid cells, possessing type I intracytoplasmic membranes and regularly arranged surface layers of linear (p2) symmetry. Strain MYHTT expressed only the particulate methane monooxygenase and employed the ribulose monophosphate pathway for formaldehyde assimilation. It is a neutrophilic and halotolerant organism capable of growth at pH 6·5–7·5 (optimum pH 6·8) and in up to 3 % NaCl (optimum 0·5–1 % NaCl). Phylogenetic analysis based on 16S rRNA gene sequence analysis indicated that strain MYHTT is most closely related to the thermophilic undescribed methanotroph ‘Methylothermus’ HB (91 % identity) and the novel halophilic methanotroph Methylohalobius crimeensis 10KiT (90 % identity). Comparative sequence analysis of particulate methane monooxygenase (pmoA) genes also confirmed the clustering of strain MYHTT with ‘Methylothermus’ HB and Methylohalobius crimeensis 10KiT (98 and 92 % derived amino acid sequence identity, respectively). The DNA G+C content was 62·5 mol%. The major cellular fatty acids were C16 : 0 (37·2 %) and C18 : 1 ω9c (35·2 %) and the major polar lipids were phosphatidylethanolamine and phosphatidylglycerol. The major ubiquinone was Q-8. On the basis of comparative phenotypic and genotypic characteristics, a new genus and species, Methylothermus thermalis gen. nov., sp. nov., is proposed, with MYHTT as the type strain (=VKM B-2345T=IPOD FERM P-19714T).

A Gram-negative, non-motile, non-spore-forming, slightly halophilic bacterial strain, SW-238T, was isolated from sea water collected from the South Sea in Korea, and subjected to a polyphasic taxonomic study. The organism grew optimally at 25–30 °C and in the presence of 2–3 % (w/v) NaCl. Strain SW-238T was characterized chemotaxonomically as containing ubiquinone (Q-8) as the predominant respiratory lipoquinone and C18 : 1 ω9c as the major fatty acid. Its DNA G+C content was 47·6 mol%. Phylogenetic analyses based on 16S rRNA gene sequences showed that strain SW-238T fell within the radiation of the cluster comprising Psychrobacter species. The 16S rRNA gene sequence of strain SW-238T had similarity levels of 94·8–97·9 % to sequences of the type strains of recognized Psychrobacter species. Levels of DNA–DNA relatedness between strain SW-238T and the type strains of 10 phylogenetically related Psychrobacter species were below 70 %. On the basis of phenotypic and phylogenetic data and genetic distinctiveness, strain SW-238T (=KCTC 12313T=JCM 12601T) was placed in the genus Psychrobacter as the type strain of a novel species, for which the name Psychrobacter celer sp. nov. is proposed.