We investigated whether ectomycorrhizal (ECM) fungal species exhibit antibacterial activity towards culturable bacterial communities in mycorrhizospheres. Four hundred and thirty bacterial strains were isolated from the ECM root tips of Pinus densiflora and bulk soil, and 21 were co-cultured with six ECM fungal species. Three hundred and twenty-nine bacterial 16S rDNA sequences were identified in ECM roots (n=185) and bulk soil (n=144). Mycorrhizosphere isolates were dominated by Gram-negative Proteobacteria from 16 genera, including Burkholderia, Collimonas, Paraburkholderia, and Rhizobium. Paraburkholderia accounted for approximately 60%. In contrast, bulk soil isolates contained a high number of Gram-positive Firmicutes, particularly from Bacillus. Paraburkholderia accounted for ≤20% of the bacterial isolates from bulk soil, which was significantly lower than its percentage in ECM root tips. Co-cultures of six ECM fungal species with the 21 bacterial strains revealed that eight strains of three Gram-positive genera—Arthrobacter, Bacillus, and Lysinibacillus—were sensitive to the antibacterial activity of the fungi. In contrast, the Gram-negative strains, including five Paraburkholderia strains, two Burkholderia strains, and a Rhizobium sp., were not sensitive. The strength of fungal antibacterial activity varied in a species-dependent manner, but consistently affected Gram-positive bacteria. These results suggest that Gram-positive bacteria are excluded from the mycorrhizosphere by the antibacterial activity of ECM fungi, which develops specific soil bacterial communities in the mycorrhizosphere.
Oil palm empty fruit bunches (OPEFB) are the most abundant, inexpensive, and environmentally friendly lignocellulosic biomass in Malaysia. Investigations on the microbial diversity of decaying OPEFB may reveal microbes with complex enzymes that have the potential to enhance the conversion of lignocellulose into second-generation biofuels as well as the production of other value-added products. In the present study, fungal and bacterial diversities in decaying OPEFB were identified using Illumina MiSeq sequencing of the V3 region of the 16S rRNA gene and V4 region of the 18S rRNA gene. Fungal diversity in decaying OPEFB was dominated by the phylum Ascomycota (14.43%), while most of the bacterial sequences retrieved belonged to Proteobacteria (76.71%). Three bacterial strains isolated from decaying OPEFB, designated as S18, S20, and S36, appeared to grow with extracted OPEFB-lignin and Kraft lignin (KL) as the sole carbon source. 16S rRNA gene sequencing identified the 3 isolates as Paenibacillus sp.. The molecular weight distribution of KL before and after degradation showed significant depolymerization when treated with bacterial strains S18, S20, and S36. The presence of low-molecular-weight lignin-related compounds, such as vanillin and 2-methoxyphenol derivatives, which were detected by a GC-MS analysis, confirmed the KL-degrading activities of isolated Paenibacillus strains.
Limited information is currently available on the assembly processes (deterministic vs. stochastic) shaping the compositions of key microbial communities in activated sludge (AS). The relative importance of deterministic and stochastic processes for key bacterial and archaeal assemblages (i.e., core-satellite and habitat generalist-specialist) in AS from 13 wastewater treatment plants in China was investigated using 16S rDNA amplicon sequencing. The results obtained indicated 1,388 and 369 core operational taxonomic units (OTUs), 1,038 and 1,683 satellite OTUs, 255 and 48 habitat generalist OTUs, and 192 and 111 habitat specialist OTUs for Bacteria and Archaea, respectively. The proportions of shared OTUs between core and habitat specialist communities were similar to or higher than those between core and habitat generalist communities, suggesting a stronger inter-linkage between the former two groups. Deterministic processes, indicated by abundance-based β-null models, were responsible for shaping core communities, in which NH4-N, OrgC/OrgN, Cr, and Ni were the main controlling factors. In contrast, satellite communities were predominantly influenced by stochastic processes. Moreover, we found that deterministic and stochastic processes were mainly responsible for shaping the assembly of habitat specialists and generalists, respectively. However, the influence of deterministic factors on habitat specialists remains unclear. The present study provides novel insights into the assembly mechanisms of AS microbial communities.
Although Pantoea species are widely distributed among plants, water, soils, humans, and animals, due to a lack of efficient isolation methods, the clonality of Pantoea species is poorly characterized. Therefore, we developed a new semi-selective medium designated ‘lysine-ornithine-mannitol-arginine-charcoal’ (LOMAC) to isolate these species. In an inclusive and exclusive study examining 94 bacterial strains, all Pantoea strains exhibited yellow colonies on LOMAC medium. The performance of the medium was assessed using Pantoea-spiked soils. Percent average agreement relative to the Api20E biochemical test was 97%. A total of 24 soil spot samples and 19 plant types were subjected to practical trials. Of the 91 yellow colonies selected on LOMAC medium, 81 were correctly identified as Pantoea species using the biochemical test. The sequencing of 16S rRNA (rrs) and gyrB from these isolates confirmed that Pantoea agglomerans, P. vagans, P. ananatis, and P. deleyi were present in Japanese fields. A phylogenetic analysis using rrs enabled only the limited separation of strains within each Pantoea spp., whereas an analysis using gyrB revealed higher variability and enabled the finer resolution of distinct branches. P. agglomerans isolates were divided into 3 groups, 2 of which were new clades, with the other comprising a large group including biocontrol strains. P. vagans was also in one of the new clades. The present results indicate that LOMAC medium is useful for screening Pantoea species. The use of LOMAC medium will provide new opportunities for identifying the beneficial properties of Japanese Pantoea isolates.
Despite efforts to address the composition of the microbial community during the anaerobic treatment of palm oil mill effluent (POME), its composition in relation to biodegradation in the full-scale treatment system has not yet been extensively examined. Therefore, a thorough analysis of bacterial and archaeal communities was performed in the present study using MiSeq sequencing at the different stages of the POME treatment, which comprised anaerobic as well as facultative anaerobic and aerobic processes, including the mixed raw effluent (MRE), mixing pond, holding tank, and final discharge phases. Based on the results obtained, the following biodegradation processes were suggested to occur at the different treatment stages: (1) Lactobacillaceae (35.9%) dominated the first stage, which contributed to high lactic acid production; (2) the higher population of Clostridiaceae in the mixing pond (47.7%) and Prevotellaceae in the holding tank (49.7%) promoted acetic acid production; (3) the aceticlastic methanogen Methanosaetaceae (0.6–0.8%) played a role in acetic acid degradation in the open digester and closed reactor for methane generation; (4) Syntrophomonas (21.5–29.2%) appeared to be involved in the degradation of fatty acids and acetic acid by syntrophic cooperation with the hydrogenotrophic methanogen, Methanobacteriaceae (0.6–1.3%); and (5) the phenols and alcohols detected in the early phases, but not in the final discharge phase, indicated the successful degradation of lignocellulosic materials. The present results contribute to a better understanding of the biodegradation mechanisms involved in the different stages of the full-scale treatment of POME.
The phylogenetic diversity and antimicrobial resistance (AMR) of Campylobacter coli from humans and animals in Japan between 2008 and 2014 were investigated. A total of 338 foodborne campylobacterioses were reported in Osaka, and C. coli was isolated from 38 cases (11.2%). In the present study, 119 C. coli strains (42 from humans, 25 each from poultry, cattle, and swine, and 2 from wild mallard) were examined by multilocus sequence typing (MLST). MLST assigned 36 sequence types (STs), including 14 novel STs; all human strains and 91% of animal strains (70/77) were assigned to the ST-828 clonal complex. The predominant human ST was ST-860 (18/42, 43%), followed by ST-1068 (8/42, 19%); these STs were also predominant in poultry (ST-860, 9/25, 36%) and cattle (ST-1068, 18/25, 72%). ST-1562 was only predominant in swine (11/25, 44.0%). Swine strains showed the greatest resistance to erythromycin (EM; 92.0%), while EM resistance was only found in 2 out of the 42 human strains examined (4.8%). All EM-resistant swine strains (n=15) exhibited a common point mutation in the 23S rRNA sequence (A2085G), and the tetO gene was detected in 22 out of the 23 TET-resistant swine strains. A whole genome sequencing analysis of four representative swine ST-1562 strains revealed abundant AMR-associated gene clusters in their genomes, suggesting horizontal gene transfer events during host adaptation. This is the first study to demonstrate the phylogenetic diversity and AMR profiles of C. coli in Japan. The present results suggest that poultry and cattle are major reservoirs, improving our knowledge on the epidemiological and ecological traits of this pathogen.
Aphids have a mutualistic relationship with the bacterial endosymbiont Buchnera aphidicola. We previously reported seven cysteine-rich peptides in the pea aphid Acyrthosiphon pisum and named them Bacteriocyte-specific Cysteine-Rich (BCR) peptides; these peptides are exclusively expressed in bacteriocytes, special aphid cells that harbor symbionts. Similar symbiotic organ-specific cysteine-rich peptides identified in the root nodules of leguminous plants are named Nodule-specific Cysteine-Rich (NCR) peptides. NCR peptides target rhizobia in the nodules and are essential for symbiotic nitrogen fixation. A BacA (membrane protein) mutant of Sinorhizobium is sensitive to NCR peptides and is unable to establish symbiosis. Based on the structural and expressional similarities between BCR peptides and NCR peptides, we hypothesized that aphid BCR peptides exhibit antimicrobial activity, similar to some NCR peptides. We herein synthesized BCR peptides and investigated their antimicrobial activities and effects on the bacterial membrane of Escherichia coli. The peptides BCR1, BCR3, BCR5, and BCR8 exhibited antimicrobial activities with increased membrane permeability. An sbmA mutant of E. coli, a homolog of bacA of S. meliloti, was more sensitive to BCR peptides than the wild type. Our results suggest that BCR peptides have properties that may be required to control the endosymbiont, similar to NCR peptides in legumes.
Chloroxylenol (CHL) is an antimicrobial ingredient that is frequently used in antiseptics/disinfectants for skin (e.g. hand soap) and non-living surfaces. CHL is an alternative to triclosan and triclocarban, the use of which has recently been banned in some countries. Accordingly, the more widespread use of CHL may significantly increase its occurrence and level in aquatic environments in the near future, eventually resulting in potential ecological risks. Wastewater treatment plants (WWTPs) may be a point source of CHL in natural environments due to extensive discharge through urban waste stream disposal. While the satisfactory removal of CHL in WWTPs is critical for maintaining healthy aquatic ecosystems, the extent of CHL removal and whether CHL causes system upset/failure in WWTPs currently remain unknown. In the present study, we conducted bioreactor operation and batch experiments to investigate the fate and effects of CHL and elucidate the mechanisms underlying degradation at various levels from environmentally relevant to high levels (0.5–5 mg L–1). Bioreactors partially removed CHL (44–87%) via a largely biological route. Microbial association networks constructed using 16S rRNA gene sequencing data revealed selective enrichment and a correlation between Sphingobium and CHL, implying its involvement in the biological breakdown of CHL through dehalogenation and ring hydroxylation pathways. The present results provide insights into the behavior and effects of CHL in activated sludge communities and important information for the sustainable management of CHL that may be an emerging issue in the urban water cycle.
The responses of bacterial communities to halocarbon were examined using a 28-d incubation of bromoform- and methanol-enriched subarctic surface seawater. Significant increases were observed in dibromomethane concentrations and bacterial 16S rRNA gene copy numbers in the treated substrates incubated for 13 d. The accumulated bacterial community was investigated by denaturing gradient gel electrophoresis and amplicon analyses. The dominant genotypes corresponded to the genera Roseobacter, Lentibacter, and Amylibacter; the family Flavobacteriaceae; and the phylum Planctomycetes, including methylotrophs of the genus Methylophaga and the family Methylophilaceae. Therefore, various phylotypes responded along with the dehalogenation processes in subarctic seawater.