Changes in ¹⁵N/¹⁴N in the soil microbial biomass during nitrogen (N) mineralization have been hypothesized to influence ¹⁵N/¹⁴N in soil organic matter among ecosystem sites. However, a direct experimental test of this mechanism has not yet been performed. To evaluate the potential control of microbial N mineralization on the natural N isotope composition, we cultured fungi (Aspergillus oryzae) in five types of media of varying C:N ratios of 5, 10, 30, 50, and 100 for 4 d, and tracked changes in δ¹⁵N in the microbial biomass, NH₄⁺, and dissolved organic N (DON: glycine) over the course of the experiment. High rates of NH₄⁺ excretion from A. oryzae were accompanied by an increase in δ¹⁵N in the microbial biomass in low C:N media (i.e., C/N<30). In contrast, NH₄⁺ was strongly retained in higher C/N treatments with only minor (i.e., <1 ‰) changes being detected in δ¹⁵N in the microbial biomass. Differences in δ¹⁵N in the microbial biomass were attributed to the loss of low-δ¹⁵N NH₄⁺ in low, but not high C/N substrates. We also detected a negative linear correlation between microbial nitrogen use efficiency (NUE) and Δ¹⁵N (δ¹⁵N-biomass–δ¹⁵N-glycine). These results suggest an isotope effect during NH₄⁺ excretion in relatively N-repleted environments in which microbial NUE is low, which may explain the vertical patterns of organic matter δ¹⁵N in soil profiles.

Nitrous oxide (N₂O) is a powerful greenhouse gas; however, limited information is currently available on the microbiomes involved in its sink and source in seagrass meadow sediments. Using laboratory incubations, a quantitative PCR (qPCR) analysis of N₂O reductase (nosZ) and ammonia monooxygenase subunit A (amoA) genes, and a metagenome analysis based on the nosZ gene, we investigated the abundance of N₂O-reducing microorganisms and ammonia-oxidizing prokaryotes as well as the community compositions of N₂O-reducing microorganisms in in situ and cultivated sediments in the non-eelgrass and eelgrass zones of Lake Akkeshi, Japan. Laboratory incubations showed that N₂O was reduced by eelgrass sediments and emitted by non-eelgrass sediments. qPCR analyses revealed that the abundance of nosZ gene clade II in both sediments before and after the incubation as higher in the eelgrass zone than in the non-eelgrass zone. In contrast, the abundance of ammonia-oxidizing archaeal amoA genes increased after incubations in the non-eelgrass zone only. Metagenome analyses of nosZ genes revealed that the lineages Dechloromonas-Magnetospirillum-Thiocapsa and Bacteroidetes (Flavobacteriia) within nosZ gene clade II were the main populations in the N₂O-reducing microbiome in the in situ sediments of eelgrass zones. Sulfur-oxidizing Gammaproteobacteria within nosZ gene clade II dominated in the lineage Dechloromonas-Magnetospirillum-Thiocapsa. Alphaproteobacteria within nosZ gene clade I were predominant in both zones. The proportions of Epsilonproteobacteria within nosZ gene clade II increased after incubations in the eelgrass zone microcosm supplemented with N₂O only. Collectively, these results suggest that the N₂O-reducing microbiome in eelgrass meadows is largely responsible for coastal N₂O mitigation.

Arbuscular mycorrhizal (AM) fungi are important members of the root microbiome and may be used as biofertilizers for sustainable agriculture. To elucidate the impact of AM fungal inoculation on indigenous root microbial communities, we used high-throughput sequencing and an analytical pipeline providing fixed operational taxonomic units (OTUs) as an output to investigate the bacterial and fungal communities of roots treated with a commercial AM fungal inoculum in six agricultural fields. AM fungal inoculation significantly influenced the root microbial community structure in all fields. Inoculation changed the abundance of indigenous AM fungi and other fungal members in a field-dependent manner. Inoculation consistently enriched several bacterial OTUs by changing the abundance of indigenous bacteria and introducing new bacteria. Some inoculum-associated bacteria closely interacted with the introduced AM fungi, some of which belonged to the genera Burkholderia, Cellulomonas, Microbacterium, Sphingomonas, and Streptomyces and may be candidate mycorrhizospheric bacteria that contribute to the establishment and/or function of the introduced AM fungi. Inoculated AM fungi also co-occurred with several indigenous bacteria with putative beneficial traits, suggesting that inoculated AM fungi may recruit specific taxa to confer better plant performance. The bacterial families Methylobacteriaceae, Acetobacteraceae, Armatimonadaceae, and Alicyclobacillaceae were consistently reduced by the inoculation, possibly due to changes in the host plant status caused by the inoculum. To the best of our knowledge, this is the first large-scale study to investigate interactions between AM fungal inoculation and indigenous root microbial communities in agricultural fields.

The Gram-negative marine propylene-assimilating bacterium, strain PE-TB08W, was isolated from surface seawater. A structural gene analysis using the 16S rRNA gene showed 96, 94, and 95% similarities to Halioglobus species, Haliea sp. ETY-M, and Haliea sp. ETY-NAG, respectively. A phylogenetic tree analysis showed that strain PE-TB08W belonged to the EG19 (Chromatocurvus)-Congregibacter-Haliea cluster within the Halieaceae (formerly Alteromonadaceae) family. Thus, strain PE-TB08W was characterized as a newly isolated Halieaceae bacterium; we suggest that this strain belongs to a new genus. Other bacterial characteristics were investigated and revealed that strain PE-TB08W assimilated propylene, n-butane, 1-butene, propanol, and 1-butanol (C3 and C4 gaseous hydrocarbons and primary alcohols), but not various other alcohols, including methane, ethane, ethylene, propane, and i-butane. The putative alkene monooxygenase (amo) gene in this strain was a soluble methane monooxygenase-type (sMMO) gene that is ubiquitous in alkene-assimilating bacteria for the initial oxidation of alkenes. In addition, two epoxide carboxylase systems containing epoxyalkane, the co-enzyme M transferase (EaCoMT) gene, and the co-enzyme M biosynthesis gene, were found in the upstream region of the sMMO gene cluster. Both of these genes were similar to those in Xanthobacter autotrophicus Py2 and were inductively expressed by propylene. These results have a significant impact on the genetic relationship between terrestrial and marine alkene-assimilating bacteria.

The climate, topography, fauna, and flora of Venezuela are highly diverse. However, limited information is currently available on the characterization of soybean rhizobia in Venezuela. To clarify the physiological and genetic diversities of soybean rhizobia in Venezuela, soybean root nodules were collected from 11 soil types located in different topographical regions. A total of 395 root nodules were collected and 120 isolates were obtained. All isolates were classified in terms of stress tolerance under different concentrations of NaCl and Al³⁺. The tolerance levels of isolates to NaCl and Al³⁺ varied. Based on sampling origins and stress tolerance levels, 44 isolates were selected for further characterization. An inoculation test indicated that all isolates showed the capacity for root nodulation on soybean. Based on multilocus sequence typing (MLST), 20 isolates were classified into the genera Rhizobium and Bradyrhizobium. The remaining 24 isolates were classified into the genus Burkholderia or Paraburkholderia. There is currently no evidence to demonstrate that the genera Burkholderia and Paraburkholderia are the predominant soybean rhizobia in agricultural fields. Of the 24 isolates classified in (Para) Burkholderia, the nodD–nodB intergenic spacer regions of 10 isolates and the nifH gene sequences of 17 isolates were closely related to the genera Rhizobium and Bradyrhizobium, respectively. The root nodulation numbers of five (Para) Burkholderia isolates were higher than those of the 20 α-rhizobia. Furthermore, among the 44 isolates tested, one Paraburkholderia isolate exhibited the highest nitrogen-fixation activity in root nodules.

Vitex rotundifolia L. f. (Lamiaceae), which commonly grows at sand coasts, is important for coast protection and the prevention of erosion. However, the diversity and roles of fungi associated with this plant remain unclear. A total of 1,052 endophytic isolates from 1,782 plants tissues from two sand beaches in northern Taiwan were classified into 76 morphospecies based on culture morphology and ITS or LSU rRNA gene sequence comparisons. Critical species were further identified using protein gene sequences and microscopy. Most of the isolates at both sites belonged to the phylum Ascomycota, with Pleosporales having the most species (15 species). The largest number of isolates (47.7%) was from the stems, followed by the roots (22.5%), leaves (16.6%), and branches (13.1%). The three species with the highest isolation frequencies at both sites were Alternaria alternata, Aspergillus terreus, and an undescribed species of Alpestrisphaeria. A. terreus was found in all organs. A. alternata was detected in all organs, except the roots. Alpestrisphaeria sp. was only found in the roots and stems. In the stems and roots, strain numbers from cortical tissues were approximately two-fold higher than those from the corresponding woody tissue. The overall colonization rate in the stems was significantly higher than those that in the roots and leaves. The majority of fungi appeared to be saprobes, which may play important roles in nutrient recycling during sand burial and mediate further stress factors in the coastal habitat.

Legumes form root nodules and fix atmospheric nitrogen by establishing symbiosis with rhizobia. However, excessive root nodules are harmful to plants because of the resulting overconsumption of energy from photosynthates. The delay of an inoculation of the soybean super-nodulation mutant NOD1–3 with Bradyrhizobium diazoefficiens USDA110^T by 5 d after an inoculation with several soil bacteria confirmed that one bacterial group significantly decreased root nodules throughout the study period. Moreover, no significant changes were observed in nitrogen fixation by root nodules between an inoculation with USDA 110^T only and co-inoculation treatments. To clarify the potential involvement of PR proteins in the restriction of nodule formation in the plants tested, the relative expression levels of PR-1, PR-2, PR-5, and PDF1.2 in NOD1–3 roots were measured using real-time PCR. One group of soil bacteria (Gr.3), which markedly reduced nodule numbers, significantly induced the expression of PR-1, PR-5 and PDF1.2 genes by day 5 after the inoculation. By days 7, 10, and 20 after the inoculation, the expression levels of PR-2 and PR-5 were lower than those with the uninoculated treatment. Inoculations with this group of soil bacteria resulted in lower root nodule numbers than with other tested soil bacteria exerting weak inhibitory effects on nodulation, and were accompanied by the induction of plant defense-related genes. Thus, PR genes appear to play important roles in the mechanisms that suppresses nodule formation on soybean roots.

The sorption and distribution of nickel, a common metal contaminant in aquatic systems, were assessed in bacterial microcolonies using a combination of fluorescent staining with Newport Green and confocal laser scanning microscopy (CLSM) with confirmation by scanning electron microscopy (SEM) and X-ray microprobe analyses. CLSM with Newport Green, selected fluor-conjugated lectins, and DNA staining allowed for the discrimination of the microdomains present in the microcolony exopolymeric matrix and detection of bound nickel. This approach avoided the artefacts associated with drying and fixation required by analytical electron microscopy. The results obtained indicated that specific microcolonies within river biofilms sorbed nickel within limited microdomains present in the complex tripartite exopolymeric matrix surrounding bacterial cells. Sorption occurred such that nickel was concentrated within the exopolymeric matrix, but not directly associated with cells. These microdomains appeared to have neutral pH and be dominated by negatively charged residues favoring the sorption of nickel and other cations. These results also suggest an important role for specific community members in the sorption and concentration of metals in aquatic biofilm communities.

Marine microbes play a central role in driving biogeochemical cycles. Microbial extracellular enzymatic activities (EEA) are the ‘gatekeeper’ of the marine carbon cycle, and these enzymes may be found attached to cells or dissolved (cell-free). Recent studies indicated that the proportion of dissolved enzymatic activity is generally similar to (if not higher than) cell-attached activity. Thus, it is critical to understand the sources and sinks of cell-free EEA in the ocean. We herein empirically tested whether bacterial stress and mortality (induced by mitomycin C) are a source of the cell-free EEA of alkaline phosphatase (APase), beta-glucosidase (BGase), and leucine aminopeptidase (LAPase). We found that bacterial stress and mortality caused relative increases in the proportion of dissolved relative to total EEA of up to 10.5% for APase, 13.5% for BGase, and 7.3% for LAPase. These relative increases in dissolved EEA corresponded to absolute increases in the cell-free pool of 4.8, 7.2, and 3.8% for APase, BGase and LAPase, respectively. Collectively, our results contribute relevant information on the origin of free dissolved extracellular enzymes in marine waters, indicating that bacterial stress and mortality are a source of cell-free enzymatic activity and suggesting a potential link between microbial interactions and the degradation of organic matter via the release of cell-free enzymes.

The present study characterized the interactions of microbial populations in activated sludge systems during the operational period after an increase in the wastewater flow rate and consequential ammonia accumulation using a 16S rRNA gene sequencing-based network analysis. Two hundred microbial populations accounting for 81.8% of the total microbiome were identified. Based on a co-occurrence analysis, Nitrosomonas-type ammonia oxidizers had one of the largest number of interactions with diverse bacteria, including a bulking-associated Thiothrix organism. These results suggest that an increased flow rate has an impact on constituents by changing ammonia concentrations and also that Nitrosomonas- and Thiothrix-centric responses are critical for ammonia removal and microbial community recovery.

Supplementation with conductive magnetite particles promoted methanogenic acetate degradation by microbial communities enriched from the production water of a high-temperature petroleum reservoir. A microbial community analysis revealed that Petrothermobacter spp. (phylum Deferribacteres), known as thermophilic Fe(III) reducers, predominated in the magnetite-supplemented enrichment, whereas other types of Fe(III) reducers, such as Thermincola spp. and Thermotoga spp., were dominant under ferrihydrite-reducing conditions. These results suggest that magnetite induced interspecies electron transfer via electric currents through conductive particles between Petrothermobacter spp. and methanogens. This is the first evidence for possible electric syntrophy in high-temperature subsurface environments.

Vibrio fischeri possesses a complex AHL-mediated Quorum-sensing (QS) system including two pathways, LuxI/R (3-oxo-C6-HSL and C6-HSL) and AinS/R (C8-HSL), which are important for the regulation of physiological traits. Diverse QS-dependent functional phenotypes have been described in V. fischeri; however, AHL diversity is still underestimated. In the present study, we investigated AHL diversity in five symbiotic V. fischeri strains with distinct phenotypic properties using UHPLC-HRMS/MS. The results obtained (1) revealed an unexpectedly high diversity of signaling molecules, (2) emphasized the complexity of QS in V. fischeri, and (3) highlight the importance of understanding the specificity of AHL-mediated QS.

Alcanivorax borkumensis is a ubiquitous marine bacterium that utilizes alkanes as a sole carbon source. We observed two phenotypes in the A. borkumensis SK2 type strain: rough (R) and smooth (S) types. The S type exhibited lower motility and higher polysaccharide production than the R type. Full genome sequencing revealed a mutation in the S type involved in cyclic-di-GMP production. The present results suggest that higher c-di-GMP levels in the S type control the biofilm forming behavior of this bacterium in a manner commensurate with other Gram-negative bacteria.

Soil bacterial community compositions and temporal changes in organic paddy fields were elucidated using a 16S rRNA gene amplicon sequencing analysis with a high-throughput next generation sequencer. At transplanting, bacterial community compositions in organic and conventional paddy fields were mostly similar despite differences in field management. The bacterial community composition in organic fields differed from that under conventional management during the rice growth period, possibly as a result of the decomposition process of organic fertilizers. However, differences in the frequency of tillage and photosynthetic bacterial inoculations in the organic plots had less of an impact on bacterial communities.