This study aimed to explore the diversity of river water Acinetobacter populations using culture-dependent and -independent methods. Pyrosequencing indicated that 1.5% of the total sequences from Qiandeng River water were classified as Acinetobacter. Twelve Acinetobacter strains were isolated from three different sampling sites of the Qiandeng River. Based on culture-dependent methods, A. johnsonii, A. lwoffii and A. guillouiae were the most abundantly represented Acinetobacter strains among the upper, middle and downstream populations of the river. Probing of three Acinetobacter-enriched 16S rRNA gene libraries with the Acinetobacter specific probe Act660F revealed 42 unique 16S rRNA gene sequences exhibiting a similarity of 94.9−99.9% with the known Acinetobacter strains. Among the uncultured Acinetobacter sequences, 50%, 58.3% and 68.8% of those obtained from upstream sampling site A, middle stream sampling site B and downstream sampling site C were phylogenetically located within Group I. This Group represented the most abundant strains of Acinetobacter populations in river water based on culture-independent methods. The results indicated that culture-independent methods provide more detailed information on the diversity of Acinetobacter populations than that based on culture-dependent methods. Therefore, the development of new and efficient isolation methods to identify uncultured Acinetobacter species is required.
The GenBank/EMBL/DDBJ accession number for the 16S rRNA gene sequence of strain DCY69T is JX233806. A Gram-reaction-negative, oxidase- and catalase-positive, non-gliding motile strain, designated strain DCY69T, was isolated from the soil of a ginseng field in the Republic of Korea. Colonies of strain DCY69T were circular, 0.5−1.5 mm diameter, yellow, and convex on an R2A agar plate after 2 days. Phylogenetic analyses based on 16S rRNA gene sequences revealed that strain DCY69T belonged to the genus Flavobacterium with 90.5−98.3% gene sequence similarity. The major predominant quinone was MK-6. The major cellular fatty acids were iso-C15:0, iso-C17:0 3-OH, iso-C15:0 3-OH and summed feature 3 (containing C16:1ω7c and/or C16:1ω6c). The major polar lipids were phosphatidylethanolamine, one unidentified aminolipid and unidentified polar lipids (L1, L2). The genomic DNA G+C content of strain DCY69T was 35.0mol%. The strain DCY69T transformed ginsenoside Rb1 into Rd and F2. Based on the polyphasic taxonomic data, strain DCY69T is considered to represent a novel species of the genus Flavobacterium, for which the name Flavobacterium panaciterrae sp. nov. is proposed. The type strain is DCY69T(= KCTC 32392T = JCM 19161T), isolated from the soil of a ginseng field in the Republic of Korea.
Dokdo, located east of the mainland of South Korea, is a volcanic island designated as a natural monument of South Korea due to its ecological value. Dokdo is divided into Dongdo and Seodo, islands with geological differences. The soil bacterial communities on Dokdo (Dongdo and Seodo) were analyzed using the pyrosequencing method. There were 1,693 and 1,408 operational taxonomic units (OTU) from Dongdo and Seodo, respectively. The statistical analyses (rarefaction curves as well as Chao1, Shannon, and Simpson indices) showed that bacterial diversity was slightly higher in Dongdo than Seodo. From results of a BLASTN search against the EzTaxon-e database, the validated reads (obtained after sequence preprocessing) were almost all classified at the phylum level. From the phylum level down to the species level, the number of classified reads considerably decreased due to the absence of information concerning unculturable or unidentified bacteria to date. Among the 36 phyla identified, three phyla (Proteobacteria, Actinobacteria and Acidobacteria) accounted for around 74.64%. The taxonomic composition was similar at the higher ranks (family and above) between Dongdo and Seodo, but a little different at the genus level. There were also various differences in the relative abundance of taxonomic ranks between Dongdo and Seodo. In particular, the proportion of the genus Acidobacterium (of the phylum Acidobacteria) was about six times higher in Seodo than Dongdo. In addition, the percentage of the genus Mycobacterium (of the phylum Actinobacteria) was nearly three times higher in Seodo than Dongdo, and the proportion of the genus Gaiella was about 3.7 times higher in Dongdo than Seodo. Overall, through the metagenomic analysis, the number of species identified in Dongdo and Seodo was 1,239 and 1,055, respectively. This information on the numerous culturable and unculturable bacteria is expected to help in the screening of new species in Dokdo.
We compared pairs of 1,226 bacterial strains with whole genome sequences and calculated their average nucleotide identity (ANI) between genomes to determine whether whole genome comparison can be directly used for bacterial species definition. We found that genome comparisons of two bacterial strains from the same species (SGC) have a significantly higher ANI than those of two strains from different species (DGC), and that the ANI between the query and the reference genomes can be used to determine whether two genomes come from the same species. Bacterial species definition based on ANI with a cut-off value of 0.92 matched well (81.5%) with the current bacterial species definition. The ANI value was shown to be consistent with the standard for traditional bacterial species definition, and it could be used in bacterial taxonomy for species definition. A new bioinformatics program (ANItools) was also provided in this study for users to obtain the ANI value of any two bacterial genome pairs (http://genome.bioinfo-icdc.org/). This program can match a query strain to all bacterial genomes, and identify the highest ANI value of the strain at the species, genus and family levels respectively, providing valuable insights for species definition.
The carbon storage regulator (Csr) global regulatory system is conserved in many eubacteria and coordinates the expression of various genes that facilitate adaptation during the major physiological growth phase. The Csr system in Escherichia coli comprises an RNA-binding protein, CsrA; small non-coding RNAs, CsrB and CsrC; and a decay factor for small RNAs, CsrD. In this study, we identified the Csr system in Serratia marcescens 2170. S. marcescens CsrA was 97% identical to E. coli CsrA. CsrB and CsrC RNAs had typical stem-loop structures, including a GGA motif that is the CsrA binding site. CsrD was composed of N-terminal two times transmembrane region and HAMP-like, GGDEF, and EAL domains. Overexpression of S. marcescens csr genes complemented the phenotype of E. coli csr mutants. S. marcescens CsrD affected the decay of CsrB and CsrC RNAs in E. coli. These resultssuggest that the Csr system in S. marcescens is composed of an RNA-binding protein, two Csr small RNAs, and a decay factor for Csr small RNAs.