Microbially produced fertilizer provides rhizobacteria to hydroponic tomato roots by forming beneficial biofilms.

Hydroponic cultivation of Solanum lycopersicum (tomato) is important, and high tomato production depends on the use of nitrogen and phosphate fertilizers. We had developed a microbial fertilizer (MF), which is mainly composed of nitrate. To investigate the effect of MF on plant growth, hydroponic tomato was grown with MF or commercial inorganic fertilizer (IF), and the microbiomes of the rhizosphere and the liquid phase were analyzed by confocal microscopy and high-throughput sequencing. Plant biomass and biofilm formation were increased by growth in MF compared to IF. The microbial community structures of tomato roots and hydroponic water differed between the two conditions, and three operational taxonomic units (OTUs) dominated in plants grown with MF. The three OTUs were related to Rudaea spp., Chitinophaga spp., and Stenotrophobacter terrae, which are reported to be disease-suppressive epiphytic or endophytic microbes of plant roots. Because these three OTUs also predominated in the MF itself, they were likely provided to the rhizosphere or endophytic environments of tomato roots via hydroponic water. KEY POINTS: • Microbial fertilizer for hydroponic growth enhanced biofilm formation on tomato root. • Microbial fertilizer contains tomato-root epiphytic or endophytic microbes. • Microbial fertilizer provided beneficial microbes to the rhizosphere and endophytic environments of tomato roots via hydroponic water.

Carotenoid Nostoxanthin Production by Sphingomonas sp. SG73 Isolated from Deep Sea Sediment.

Carotenoids are used commercially for dietary supplements, cosmetics, and pharmaceuticals because of their antioxidant activity. In this study, colored microorganisms were isolated from deep sea sediment that had been collected from Suruga Bay, Shizuoka, Japan. One strain was found to be a pure yellow carotenoid producer, and the strain was identified as Sphingomonas sp. (Proteobacteria) by 16S rRNA gene sequence analysis; members of this genus are commonly isolated from air, the human body, and marine environments. The carotenoid was identified as nostoxanthin ((2,3,2',3')-ß,ß-carotene-2,3,2',3'-tetrol) by mass spectrometry (MS), MS/MS, and ultraviolet-visible absorption spectroscopy (UV-Vis). Nostoxanthin is a poly-hydroxy yellow carotenoid isolated from some photosynthetic bacteria, including some species of Cyanobacteria. The strain Sphingomonas sp. SG73 produced highly pure nostoxanthin of approximately 97% (area%) of the total carotenoid production, and the strain was halophilic and tolerant to 1.5-fold higher salt concentration as compared with seawater. When grown in 1.8% artificial sea salt, nostoxanthin production increased by 2.5-fold as compared with production without artificial sea salt. These results indicate that Sphingomonas sp. SG73 is an efficient producer of nostoxanthin, and the strain is ideal for carotenoid production using marine water because of its compatibility with sea salt.

Efficient conversion of organic nitrogenous wastewater to nitrate solution driven by comammox Nitrospira.

A bacterium capable of complete ammonia oxidation (comammox) has been widely found in various environments, whereas its industrial application is limited due to the difficulty of cultivation and/or enrichment. We developed a biological system to produce a high-quality nitrate solution for use in hydroponic fertilizer. The system was composed of two separate reactors for ammonification and nitrification and was found to have a stable and efficient performance in the conversion of organic nitrogen to nitrate. To determine the key microbes involved and better understand the system, the microbial communities in the reactors were analyzed by 16S rRNA gene sequencing in combination with a shotgun metagenomic analysis. Canonical ammonia-oxidizing bacteria, which can only catalyze the oxidation of ammonia to nitrite, were detected with negligible relative abundances, while a comammox Nitrospira-related operational taxonomic unit (OTU) dominated the nitrification reactor. Furthermore, the comammox-type ammonia monooxygenase was found to be 500 times more highly expressed than the canonical one by quantitative PCR, indicating that comammox was the main driver of the stable and efficient ammonia oxidation in the system. A microbial co-occurrence analysis revealed a strong positive correlation between Nitrospira and several OTUs, some of which, such as Anaerolinea OTU, have been found to co-exist with comammox Nitrospira in the biofilms of water treatment systems. Given that these OTUs were abundant only on microbe-attached carriers in the system, their co-existence within the biofilm could be beneficial to stabilize the Nitrospira abundance, possibly by physically preventing oxygen exposure as well as cell spillage.

Development of an anti-norovirus single-chain Fv for immunochromatographic test kit.

It was developed that an anti-norovirus single-chain Fv (scFv) antibody for immunochromathographic test kit by using the phage displayed technique and biopanning. To obtain the scFv having wide reactivity for several norovirus genotypes, a mixture of recombinant norovirus capsid proteins, including 11 norovirus genotypes, was used for biopanning. Then, one anti-norovirus scFv antibody that recognized both of norovirus genogroups GI and GII was identified. An immunochromatographic test strip was constructed by using the scFv and demonstrated it to detect norovirus in stool samples. The immunochromatographic strip showed similar sensitivity to a commercially available one on which several monoclonal antibodies are included.

Quick selection of a chimeric T2 phage that displays active enzyme on the viral capsid.

We designed a bacteriophage T2 system to display proteins fused at the N-terminus of the head protein small outer capsid (SOC) of a T2 phage. To facilitate selection of chimeric phage, a T2 phage encoding the beta-galactosidase gene (betagal) upstream of the soc gene was constructed. The phage, named T2betaGal, produces blue plaques on agar plates containing XGal. Subsequently, a plasmid encoding the target protein upstream of soc was constructed and used to transform E. coli B(E) cells. Transformed cells were infected with T2betaGal and homologous recombination between phage DNA and the plasmid resulted in a chimeric phage that produced transparent plaques due to the excision of the betagal gene. Chitosanase of Bacillus sp. strain K17 (ChoK), consisting of 453 amino acids, was used as a model target protein. Recombinant T2 phage that produced ChoK was named T2ChoK. T2ChoK was produced from T2betaGal at a recombination frequency of about 0.1%. On the other hand, the value for T2betaGal produced from wild-type T2 was 0.001 %. This new system enables us to select recombinant phage very quickly and accurately. The number of molecules of ChoK was calculated at 14.7 per single phage. Latent period and burst size were estimated for the chimeric phages.