Laboratory-scale acidophilic nitrifying sequencing-batch reactors (ANSBRs) were constructed by seeding with sewage-activated sludge and cultivating with ammonium-containing acidic mineral medium (pH 4.0) with or without a trace amount of yeast extract. In every batch cycle, the pH varied between 2.7 and 4.0, and ammonium was completely converted to nitrate. Attempts to detect nitrifying functional genes in the fully acclimated ANSBRs by PCR with previously designed primers mostly gave negative results. 16S rRNA gene-targeted PCR and a subsequent denaturating gradient gel electrophoresis analysis revealed that a marked change occurred in the bacterial community during the overall period of operation, in which members of the candidate phylum TM7 and the class Gammaproteobacteria became predominant at the fully acclimated stage. This result was fully supported by a 16S rRNA gene clone library analysis, as the major phylogenetic groups of clones detected (>5% of the total) were TM7 (33%), Gammaproteobacteria (37%), Actinobacteria (10%), and Alphaproteobacteria (8%). Fluorescence in situ hybridization with specific probes also demonstrated the prevalence of TM7 bacteria and Gammaproteobacteria. These results suggest that previously unknown nitrifying microorganisms may play a major role in ANSBRs; however, the ecophysiological significance of the TM7 bacteria predominating in this process remains unclear.

Three actinomycetes, designated strains VN05A0342, VN05A0351 and VN05A0415 T , were isolated from plant-litter samples collected in the north of Vietnam and examined in a polyphasic taxonomic study. Phylogenetic analysis based on the 16S rRNA gene sequences showed that these isolates were most closely related to the type strain of Kineosporia mikuniensis (98.5 % sequence similarity). Morphological properties (the formation of spore domes and motile spores) and chemotaxonomic data supported the assignment of the three isolates to the genus Kineosporia. The isolates all contained the following: meso-diaminopimelic acid in the peptidoglycan (with small amounts of the LL isomer); ribose, mannose, galactose and glucose as the whole-cell sugars; MK-9(H 4 ) as the predominant isoprenoid quinone; C 18 : 1 and C 16 : 0 as the major cellular fatty acids; and phosphatidylcholine, phosphatidylglycerol, diphosphatidylglycerol and phosphatidylinositol as the phospholipids. The high DNA-DNA relatedness (.71 %) among the three isolates showed that they represented a single species. On the other hand, the DNA-DNA relatedness between the novel isolates and all type strains of Kineosporia species was less than 46 %. The physiological properties of our isolates were distinct from those of all of the Kineosporia species with validly published names, e.g. decomposition of L-tyrosine and aesculin and the utilization of raffinose and D-arabitol. Therefore, strains VN05A0342, VN05A0351 and VN05A0415T represent a novel species of the genus Kineosporia, for which the name Kineosporia babensis sp. nov. is proposed. The type strain is VN05A0415 T

A novel actinomycete, designated strain VN05A0561T , was isolated from plant litter collected at Ba Be National Park, Vietnam. The substrate mycelia and spore chains fragmented in a manner similar to nocardioform actinomycetes; the spores had smooth surfaces and were rodshaped. Strain VN05A0561 T had the following chemotaxonomic characteristics: mesodiaminopimelic acid in the cell-wall peptidoglycan, arabinose and galactose as characteristic sugars, MK-8(H 4 ) as the major isoprenoid quinone, phosphatidylcholine as the diagnostic phospholipid and iso-C 16 : 0 as the major cellular fatty acid. Strain VN05A0561 T shared low levels of 16S rRNA gene sequence similarity (,97 %) with the type strains of recognized species of the genus Pseudonocardia and could be differentiated from its closest phylogenetic relatives based on phenotypic characteristics. These results suggested that strain VN05A0561 T represents a novel species of the genus Pseudonocardia, for which the name Pseudonocardia babensis sp. nov. is proposed. The type strain is VN05A0561 T (5VTCC-A-The genus Pseudonocardia was established as a nocardioform actinomycete by Henssen (1957), and its description has been emended based on both chemotaxonomic and morphological variations (Reichert et al., 1998;Huang et al., 2002;Park et al., 2008). On the basis of the lack of mycolic acids and the presence of cell-wall type IV, the genera Amycolata (Lechevalier et al., 1986) and Pseudoamycolata were also proposed as nocardioform actinomycetes. However, the latter two genera were found to have chemotaxonomic properties similar to those of the genus Pseudonocardia (Kothe et al., 1989;Takeuchi et al., 1992), and were incorporated in the genus Pseudonocardia on the basis of 16S rRNA gene sequence analysis (Warwick et al., 1994;McVeigh et al., 1994). Subsequently, members of the genus Actinobispora (Jiang et al., 1991) were also transferred to the genus Pseudonocardia based on identification with the use of specific PCR primers, and re-analysis of 16S rRNA gene sequences and menaquinones of the type species (Huang et al., 2002 Reichert et al., 1998;Gu et al., 2006; Chen et al., 2009).In the present study, a Pseudonocardia-like strain isolated from a plant litter sample collected during the course of a study on the diversity of actinomycetes inhabiting Vietnam (Sakiyama et al., 2009) was studied by using a polyphasic taxonomic approach.Plant litter samples were collected at the Ba Be National Park, Bac Kan, in the mountainous area of northern Vietnam. Samples were dried at room temperature for 5-7 days and subsequently used for isolation. Rehydration and centrifugation methods (Hayakawa et al., 2000) were employed for isolation by using humic acid-vitamin agar (Hayakawa & Nonomura, 1987) containing nalidixic acid (20 mg l 21) and kabicidine (0.75 mg l 21). Strain VN05A0561 T was isolated after incubation for more than 10 days at room temperature. Strain VN05A0561T was cultured on yeast extract-soluble starch medium (YS medium; per litre distilled water: 2 g yeast extract...

Strain VN07A0015 T was isolated from soil collected on Cat Ba Island, Vietnam. The taxonomic position of strain VN07A0015 T was near Streptomyces aomiensis M24DS4 T (98.5 % 16S rRNA gene sequence similarity) and Streptomyces scabrisporus NBRC 100760 T (95.6 %), and it clustered within them; however, this cluster was distant from the type strains of other species of the genus Streptomyces. The aerial mycelia of strain VN07A0015 T were greyish and formed imperfect spiral spore chains (retinaculiaperti type) with smooth-surfaced spores. The morphological features of strain VN07A0015 T were different from those of the type strains of S. aomiensis and S. scabrisporus. The chemotaxonomic characteristics of strain VN07A0015 T were typical for all members of the genus Streptomyces, which possessed LL-type diaminopimelic acid, menaquinone MK-9(H 6 , H 8 ) and the major fatty acids iso-C 16 : 0 and iso-C 15 : 0 . DNA-DNA relatedness between strain VN07A0015 T and S. aomiensis NBRC 106164 T was less than 30 %. In addition, some physiological and biochemical traits differed from those of S. aomiensis. Therefore, we propose that strain VN07A0015 T be classified in the genus Streptomyces as a representative of Streptomyces catbensis sp. nov. (type strain VN07A0015 T 5VTCC-A-1889 T 5NBRC 107860 T ).

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