BackgroundThe pure culture of prokaryotes remains essential to elucidating the role of these organisms. Scientists have reasoned that hard to cultivate microorganisms might grow in pure culture if provided with the chemical components of their natural environment. However, most microbial species in the biosphere that would otherwise be “culturable” may fail to grow because of their growth state in nature, such as dormancy. That means even if scientist would provide microorganisms with the natural environment, such dormant microorganisms probably still remain in a dormant state.ResultsWe constructed an enrichment culture system for high-efficiency isolation of uncultured strains from marine sediment. Degree of enrichment analysis, dormant and active taxa calculation, viable but non-culturable bacteria resuscitation analysis, combined with metatranscriptomic and comparative genomic analyses of the interactions between microbial communications during enrichment culture showed that the so-called enrichment method could culture the “uncultured” not only through enriching the abundance of “uncultured,” but also through the resuscitation mechanism. In addition, the enrichment culture was a complicated mixed culture system, which contains the competition, cooperation, or coordination among bacterial communities, compared with pure cultures.ConclusionsConsidering that cultivation techniques must evolve further—from axenic to mixed cultures—for us to fully understand the microbial world, we should redevelop an understanding of the classic enrichment culture method. Enrichment culture methods can be developed and used to construct a model for analyzing mixed cultures and exploring microbial dark matter. This study provides a new train of thought to mining marine microbial dark matter based on mixed cultures.Electronic supplementary materialThe online version of this article (10.1186/s40168-018-0613-2) contains supplementary material, which is available to authorized users.

The taxonomic characteristics of two bacterial strains, FH5T and SS4, isolated from enrichment cultures obtained from two distinct marine environments, were determined. These bacteria were Gram-stain-negative, facultatively anaerobic rods. Growth occurred at 20–40 °C (optimum, 28–32 °C), pH 5.5–9.0 (optimum, pH 7.0–7.5) and in the presence of 1–7 % NaCl (optimum, 2–4 %). The major cellular fatty acids were anteiso-C15 : 0 and iso-C15 : 0. Menaquinone 7 (MK-7) was the sole respiratory quinone. The major polar lipids were phosphatidylethanolamine, an unkown phospholipid and an unknown lipid. The DNA G+C contents of strains FH5T and SS4 were both determined to be 42.0 mol%. The results of DNA–DNA hybridization studies indicated that the FH5T and SS4 genomes share greater than 95 % relatedness. The strains formed a distinct phyletic line within the class Bacteroidia , with less than 89.4 % sequence similarity to their closest relatives with validly published names. On the basis of physiological and biochemical characteristics, 16S rRNA gene sequences and chemical properties, a novel genus and species, Draconibacterium orientale gen. nov., sp. nov., within the class Bacteroidia , are proposed, with strain FH5T ( = DSM 25947T = CICC 10585T) as the type strain. In addition, a new family, Draconibacteriaceae fam. nov., is proposed to accommodate Draconibacterium gen. nov.

A Gram-stain-negative, facultatively anaerobic, gliding, yellow-pigmented bacterium, designated SS12T, was isolated from shark gill homogenate and characterized using a polyphasic approach. The strain was catalase-positive and oxidase-negative. Optimal growth occurred at 28–30 °C, pH 7.0–7.5 and in the presence of 2–4 % (w/v) NaCl. The DNA G+C content was 40.0 mol%. The strain contained MK-7 as the prevailing menaquinone; iso-C15 : 0 and anteiso-C15 : 0 as the major cellular fatty acids; and phosphatidylethanolamine and an unknown lipid as the predominant polar lipids. Comparative analysis of 16S rRNA gene sequences demonstrated that the novel isolate showed the highest sequence similarity (94.68 %) to Saccharicrinis fermentans DSM 9555T and the sequence similarities among the type strains of all other species studied were less than 92 %. A phylogenetic tree, based on 16S rRNA gene sequences, showed that strain SS12T and Saccharicrinis fermentans DSM 9555T formed a distinct cluster within the family Marinilabiliaceae . On the basis of its phylogenetic position and phenotypic traits, strain SS12T represents a novel species of genus Saccharicrinis , for which the name Saccharicrinis carchari sp. nov. is proposed. The type strain is SS12T ( = CICC 10590T = DSM 27040T). Emended descriptions of the genus Saccharicrinis and Saccharicrinis fermentans are also provided.

Woeseia oceani gen. nov., sp. nov., a chemoheterotrophic member of the order Chromatiales, and proposal of Woeseiaceae fam. nov. A novel Gram-stain-negative, rods or bent rods, facultatively anaerobic, oxidase-negative and catalase-positive bacterium, designated XK5 T , was isolated from coastal sediment from Xiaoshi Island, Weihai, China. Optimal growth occurred at 28-35 8C (range 8-42 8C) and pH 7.0-8.0 (range pH 6.0-9.0) with 1-3 % (w/v) NaCl (range 0.5-8 %). Phylogenetic analysis based on 16S rRNA gene sequences revealed that strain XK5 T was 92.1 % similar to the type strain of Thioalkalivibrio thiocyanodenitrificans, 91.9 % to the type strain of Thioalkalivibrio sulfidiphilus and 91.8 % to the type strain of Thioalkalivibrio denitrificans; similarity to other species was less than 91 %. The isolate and closely related environmental clones formed a novel family level clade in the order Chromatiales. The polar lipid profile of the novel isolate consisted of phosphatidylethanolamine, phosphatidylglycerol and some other unknown phospholipids, aminolipids and lipids. Major cellular fatty acids were iso-C 17 : 1 v9c and iso-C 15 : 0 and the main respiratory lipoquinone was Q-8. The DNA G+C content of strain XK5 T was 59.3 mol%. Comparative analysis of 16S rRNA gene sequences and characterization indicated that strain XK5 T represents a novel species of a new genus within a novel family of the order Chromatiales, for which the name Woeseia oceani gen. nov., sp. nov. is proposed. The type strain of Woeseia oceani is XK5 T (5ATCC BAA-2615 T 5CICC 10905 T ). In addition, a novel family name, Woeseiaceae fam. nov., is proposed to accommodate the genus Woeseia.In the past, the taxonomy of bacteria was entirely based on simple phenotypic characteristics (Imhoff, 2005), which aroused a long-time dispute about the taxonomic position of the bacteria now classified as the Ectothiorhodospiraceae since their discovery by Pelsh (1936). Members of the Ectothiorhodospiraceae were previously treated as a genus of the Chromatiaceae. Until recently, with the application of phylogenetic taxonomy of 16S rRNA, the Ectothiorhodospiraceae was separated as a family distinct from the family Chromatiaceae (Imhoff, 1984). At the time of writing, the order Chromatiales contains five families, according to the List of Prokaryotic Names with Standing in Nomenclature (LPSN; Euzéby, 1997), and members of the order are famous for their various types of metabolism, such as photoautotrophic, photoheterotrophic, chemoautotrophic and chemoheterotrophic. Most phototrophic bacteria in the order Chromatiales can utilize reduced sulfur compounds as electron donors, which forms highly refractile globules of elemental sulfur; the family Chromatiaceae deposit the sulfur globules inside the cells while the Ectothiorhodospiraceae outside (Pfennig & Trüper, 1974). In this study, a novel chemoheterotrophic bacterium that had significant differences with the members of the Ectothiorhodospiraceae, the closest relatives of the novel isolate, is reported. Based o...

Streptococcus agalactiae (Group B streptococcus) has emerged as an important pathogen that affects humans and animals, including aquatic species. S. agalactiae infections are becoming an increasing problem in aquaculture and have been reported worldwide in a variety of fish species, especially those living in warm water. Recently, a very serious infectious disease of unknown aetiology broke out in ya-fish (Schizothorax prenanti) farms in Sichuan Province. A Gram-positive, chain-forming coccus was isolated from moribund cultured ya-fish. The goals of this study were to identify the bacterial strains isolated from diseased fish between 2009 and 2011 in Sichuan Province, China, to evaluate the pathogenicity of the pathogen in ya-fish, crucian carp (Carassius carassius) and the Nile tilapia (Oreochromis niloticus); and to determine the susceptibility of the pathogen strains to many currently available anti-microbial agents. The virulence tests were conducted by intraperitoneal injection of bacterial suspensions. In this study, four strains of a Gram-positive, chain-forming coccus were isolated from moribund cultured ya-fish (S. prenanti). The coccoid microorganism was identified as S. agalactiae using a commercial streptococcal grouping kit and 16S rDNA sequencing analysis. Susceptibility of the isolates to 22 antibiotics was tested using the disc diffusion method. All isolates showed a similar antibiotic susceptibility, which were sensitive to amoxicillin, ciprofloxacin, lomefloxacin, chloramphenicol, rifampin, vancomycin, azithromycin, florfenicol, cefalexin, cefradine and deoxycycline and resistant to gentamicin, sinomin (SMZ/TMP), penicillin, tenemycin, fradiomycin and streptomycin. Furthermore, the virulence tests were conducted by intraperitoneal injection of the isolated strain GY101 in ya-fish, crucian carp and the Nile tilapia. This coccus was lethal to ya-fish, Nile tilapia and crucian carp. The mortality rates of infected ya-fish were 100%, 100%, 60% and 20% at doses of 1.0 × 10⁷, 1.0 × 10⁶, 1.0 × 10⁵ and 1.0 × 10⁴ CFU/fish, respectively. The values were 100%, 80%, 60% and 30% for Nile tilapia and 100%, 60%, 10% and 0% for crucian carp. To our knowledge, this is the first report of S. agalactiae as a pathogen of ya-fish.