Antibiotic resistance genes (ARGs) have been emerging as a concerning threat to both environment and public health. The continuous input of manure, irrigation water, and fertilizers increases the abundance of ARGs in agricultural environments. However, current risk assessments have focused on clinical settings, which are not applicable to environmental settings. Therefore, we herein aimed to identify and assess indicator genes to reduce the time and effort required for ARG surveillance. A nationwide ana­lysis of 322 ARGs and 58 mobile genetic elements (MGEs) was performed on 42 greenhouse and 19 control soil samples. The chemical properties and pH of soil were also investigated to characterize differences between greenhouse and control soil samples. The results obtained showed that the abundance of ARGS was significantly higher and ion concentrations were higher in greenhouse samples than in control samples. These results indicate that agricultural activities increased the abundance of ARGs. Furthermore, the abundance of core genes was significantly higher in greenhouse samples than in control samples, and the chemical characteristics of soil significantly differed between these samples. Among the discriminatory genes selected, tetM was identified as an ARG surveillance indicator gene based on its clinical relevance, prevalence in the soil resistome, and relationship with mobile genetic elements. The present results will contribute to the continuous and rapid surveillance of antibiotic resistance dissemination and proliferation in greenhouses in South Korea.

To enhance the growth of the cyanobacterium Synechococcus elongatus, the present study conducted direct screening for cyanobacterium growth-promoting bacteria (CGPB) using co-cultures. Of the 144 strains obtained, four novel CGPB strains were isolated and phylogenetically identified: Rhodococcus sp. AF2108, Ancylobacter sp. GA1226, Xanthobacter sp. AF2111, and Shewanella sp. OR151. A co-culture of S. elongatus with the most effective CGPB strain, Rhodococcus sp. AF2108, achieved a 8.5-fold increase in the chlo­rophyll content of cyanobacterial cells over that in a monoculture. A flow cytometric ana­lysis showed a 3.9-fold increase in the number of S. elongatus cells in the co-culture with Rhodococcus sp. AF2108. These results were attributed to increases in forward scattering and chlo­rophyll fluorescence intensities. The new Rhodococcus strain appears to be one of the most effective CGPBs described to date.

Beneficial root endophytic fungi induce systemic responses, growth promotion, and induced systemic resistance (ISR) in colonized host plants. The soil application of chitin, a main component of fungal cell walls, also systemically induces disease resistance. Therefore, chitin recognition and its downstream signaling pathway mediate ISR triggered by beneficial fungi colonizing the root. The present study compared systemic disease resistance and transcriptional responses induced by Trichoderma, a representative beneficial root endophytic fungus, and chitin in Arabidopsis. Significant plant growth promotion was observed under root colonization by the three beneficial fungi tested: Trichoderma atroviride, Serendipita indica, and S. vermifera. Only T. atroviride and S. indica triggered ISR against the necrotrophic fungal pathogen Alternaria brassicicola. Induced systemic resistance triggered by T. atroviride was compromised in the chitin-receptor mutant, whereas systemic resistance caused by the soil application of chitin was not. A transcriptome ana­lysis demonstrated that chitin-regulated genes were mostly shared with those regulated by T. atroviride; however, many of the latter were specific. The commonly enriched gene ontologies for these genes indicated that the T. atroviride inoculation and chitin application systemically controlled similar transcriptional responses, mainly associated with cell wall functions. Therefore, Trichoderma may trigger ISR primarily independent of the chitin-mediated signaling pathway; however, chitin and Trichoderma may systemically induce similar cellular functions aboveground.

Myxobacteria, belonging to the phylum Myxococcota, are ubiquitous in soil, marine, and other environments. A recent metagenomic sequencing ana­lysis showed that Myxococcota are predominant in activated sludge systems; however, their metabolic traits remain unclear. In the present study, we exami­ned the potential biological functions of 46 metagenomic bins of Myxococcota reconstructed from activated sludge samples from four municipal sewage treatment plants. The results obtained showed that most Myxococcota bins had an almost complete set of genes associated with glycolysis and the TCA cycle. The Palsa-1104 and Polyangiales bins contained the glycoside hydrolase GH5 and peptidase M23, which are presumably involved in lysis of the cell wall and cellular cytoplasm, suggesting that some Myxococcota from activated sludge prey on other microorganisms. The cell contact-dependent predatory functions of Myxococcus xanthus are conserved in the family Myxococcaceae, but not in other families. Two bins belonging to Palsa-1104 had phototrophic gene clusters, indicating the potential for heterotrophic and autotrophic metabolism by these microbes. In assessments of the social behavior of Myxococcota in activated sludge, the FruA gene and C-signal gene, which are involved in the regulation of fruiting body formation, were lacking in Myxococcota bins, suggesting their inability to form fruiting bodies. In addition, multiple bins of Myxococcota had novel secondary metabolite biosynthesis gene clusters that may be used for the predation of other bacteria in activated sludge. Our metagenome-based ana­lyses provide novel insights into the microbial interactions associated with Myxococcota in activated sludge ecosystems.

We herein investigated the effects of salt (NaCl) stress on soybean nodulation by rhizobial strains. We specifically exami­ned: (1) the effects of NaCl on nodule maturity and positioning by inoculating three rhizobial strains (Bradyrhizobium diazoefficiens USDA110^T, Bradyrhizobium elkanii USDA31, and Sinorhizobium fredii USDA191) onto soybean variety CNS, (2) the effects of the NaCl treatment on isoflavones (daidzein and genistein) secretion by CNS, (3) the effects of the NaCl treatment on gene expression induced by daidzein and genistein in rhizobia, and (4) the effects of the NaCl treatment on rhizobial growth. The results obtained were as follows: (1) the NaCl treatment delayed nodule development and reduced nodulation on the primary root following the USDA110^T inoculation, minimal sensitivity regarding nodule formation in the USDA 31 inoculation, and significantly increased the mature nodule number and nodules on the primary root following the USDA 191 inoculation. (2) The NaCl treatment significantly reduced the secretion of daidzein from soybean roots, but did not significantly affect that of genistein. (3) NaCl treatment induced a significant decrease in genistein-induced nodC expression in USDA110^T, but not in USDA31, and also caused a significant reduction in daidzein-induced nodC expression, but not genistein-induced expression, in USDA191. (4) NaCl treatment reduced survivability under acidic conditions, but increased survivability under saline-alkaline conditions for USDA191 than bradyrhizobia. These results indicate that saline conditions give S. fredii a competitive advantage over Bradyrhizobium during soybean infection.

White root rot disease caused by Rosellinia necatrix is a growing issue in orchards, and biochar pyrolyzed from the pruned branch residues of fruit trees has potential as a soil amendment agent with a number of benefits, such as long-term carbon sequestration. However, the effects of pruned branch biochar on white root rot disease remain unclear. Therefore, we compared direct antagonism against R. necatrix between soils with and without pruned pear branch biochar using a toothpick method and then linked soil physicochemical properties and microbial communities with soil antagonism. The results obtained showed that soil antagonism against the pathogen, that is, the extinction zone of R. necatrix in mycelial toothpicks, decreased in soils amended with 20% (v/v) pruned branch biochar. Soil pH was neutralized and aeration was promoted by the biochar amendment, which may be favorable for pathogen growth. An investigation of microbial communities surrounding R. necatrix mycelia indicated that antagonistic fungi affiliated with Chaetomiaceae and Trichoderma were selectively excluded from the mycosphere community in biochar-amended soil. Therefore, the enrichment of these indigenous antagonistic fungi may be important for controlling R. necatrix. Based on the present results, we do not recommend the application of pruned branch biochar to the soil area associated with the roots of fruit trees in order to avoid increasing the risk of white root rot in orchards.

Biogas digestive effluent (BDE) has been applied to rice fields in the Vietnamese Mekong Delta (VMD). However, limited information is available on the community composition and isolation of methanotrophs in these fields. Therefore, the present study aimed (i) to clarify the responses of the methanotrophic community in paddy fields fertilized with BDE or synthetic fertilizer (SF) and (ii) to isolate methanotrophs from these fields. Methanotrophic communities were detected in rhizospheric soil at the rice ripening stage throughout 2 cropping seasons, winter-spring (dry) and summer-autumn (wet). Methanotrophs were isolated from dry-season soil samples. Although the continued application of BDE markedly reduced net methane oxidation potential and the copy number of pmoA genes, a dissimilarity ordination ana­lysis revealed no significant difference in the methanotrophic community between BDE and SF fields (P=0.167). Eleven methanotrophic genera were identified in the methanotrophic community, and Methylosinus and Methylomicrobium were the most abundant, accounting for 32.3–36.7 and 45.7–47.3%, respectively. Type-I methanotrophs (69.4–73.7%) were more abundant than type-II methanotrophs (26.3–30.6%). Six methanotrophic strains belonging to 3 genera were successfully isolated, which included type I (Methylococcus sp. strain BE1 and Methylococcus sp. strain SF3) and type II (Methylocystis sp. strain BE2, Methylosinus sp. strain SF1, Methylosinus sp. strain SF2, and Methylosinus sp. strain SF4). This is the first study to examine the methanotrophic community structure in and isolate several methanotrophic strains from BDE-fertilized fields in VMD.