Characterisation and identification of carbofuran-utilising bacteria isolated from agricultural soil

A total of 96 bacterial cultures were isolated from soil. Seventeen bacterial isolates were selected following their cultivation on solid media containing 100 mg · L^?1 carbofuran as the sole source of carbon and nitrogen. Of the 17 isolates, 10F, 11M, 17N, 23B and 26M were specifically chosen because of their relatively higher growth efficiency and genetic diversity based on Box-polymerase chain reaction analysis. These bacterial cultures had 16S rRNA gene sequences that were most similar to Acinetobacter baumannii (10F), Agrobacterium tumefaciens (11M), Ochrobactrum anthropi (17N), Escherichia coli (23B) and Agrobacterium tumefaciens (26M) with 97, 95, 93, 95 and 94% similarity in their 16S rDNA gene sequence, respectively. Degradation rates of carbofuran in soil inoculated with these isolates were 1.9, 1.5, 1.6, 1.7 and 1.6 times, respectively, faster in comparison with uninoculated soil after 10 days of incubation. The maximum degradation rates of carbofuran (45 and 91%) were detected in soil inoculated with A. baumannii (10F) after 10 and 20 days’ incubation, respectively. These data indicate that these isolates may have the potential for use in bioremediation of pesticide contaminated soil.     

Isolation and characterisation of a carbofuran degrading Burkholderia sp. PCL3 from carbofuran-phytoremediated rhizosphere soil

A bacterial strain capable of degrading carbofuran as the sole carbon source was isolated from carbofuran-phytoremediated rhizosphere soil of rice. A 16S rRNA study identified the strain as Burkholderia sp. (isolate PCL3). Free cells of isolate PCL3 possessed inhibitory-type degradation kinetics with a q _max of 0.087 day^?1 and S _m of 248.76 mg·L^?1. Immobilised PCL3 on corncob and sugarcane bagasse possessed Monod-type degradation kinetics with a q _max of 0.124 and 0.098 day^?1, respectively. The optimal pH and temperature with the highest degradation rate coefficient of carbofuran were pH 7.5 and 35 °C, respectively.     

Arsenite oxidation by <Emphasis Type="Italic">Alcaligenes</Emphasis> sp. strain RS-19 isolated from arsenic-contaminated mines in the Republic of Korea

Arsenite [As(III)]-oxidizing bacteria play important roles in reducing arsenic [As] toxicity and mobility in As-contaminated areas. As-resistant bacteria were isolated from the soils of two abandoned mines in the Republic of Korea. The isolated bacteria showed relatively high resistances to As(III) up to 26 mM. The PCR-based 16S rRNA analysis revealed that the isolated As-resistant bacteria were close relatives to Serratia marcescensa, Pseudomonas putida, Pantoea agglomerans, and Alcaligenes sp. Among the five As-resistant bacterial isolates, Alcaligenes sp. strain RS-19 showed the highest As(III)-oxidizing activity in batch tests, completely oxidizing 1 mM of As(III) to As(V) within 40 h during heterotrophic growth. This study suggests that the indigenous bacteria have evolved to retain the ability to resist toxic As in the As-contaminated environments and moreover to convert the species to a less toxic form [e.g., from As(III) to As(V)] and also contribute the biogeochemical cycling of As by being involved in speciation of As.     

Biodegradation of Hexachlorocyclohexane by two species of Bacillus isolated from contaminated soil

Biodegradation of α, β, γ and δ hexachlorocyclohexane (HCH) isomers was studied in broth medium and soil microcosm by Bacillus circulans and Bacillus brevis isolated from contaminated soil. Degradation of α and γ isomers by both the bacterial isolates was higher than thermodynamically stable β and δ isomers. However, B. circulans was found more effective than B. brevis for β and δ isomers. Maximum rate of degradation was recorded at 150 mg/L followed by 100 and 50 mg/L. Soil microcosm study revealed maximum degradation of HCH isomers in the treatment containing natural soil, pesticide and bacterial inocula than the treatment having sterilised soil, pesticide and bacterial isolates. Chloride release was positively co-related with HCH degradation in broth medium as well as in soil microcosm, suggesting that B. circulans and B. brevis hold promising potential by having efficient enzyme(s) required for dechlorination of HCH from contaminated sites.     

Culturable actinobacteria isolated from marine sponge <Emphasis Type="Italic">Iotrochota</Emphasis> sp.

The study describes the diversity of actinobacteria isolated from the marine sponge Iotrochota sp. collected in the South China Sea. Species and natural product diversity of isolates were analyzed, including screening for genes encoding polyketide synthases (PKS) and nonribosomal peptide synthetase (NRPS), and 16S rRNA gene restriction fragment length polymorphism (RFLP). PKS and NRPS sequences were detected in more than half of the isolates and the different “PKS-I–PKS-II–NRPS” combinations in different isolates belonging to the same species indicated a potential natural product diversity and divergent genetic evolution. The phylogenetic analysis based on 16S rRNA gene sequencing showed that the isolates belonged to genera Streptomyces, Cellulosimicrobium, and Nocardiopsis. The majority of the strains tested belonged to the genus Streptomyces and one of them may be a new species. To our knowledge, this is the first report of a bacterium classified as Cellulosimicrobium sp. isolated from a marine sponge. Key Laboratory of Marine Bio-recourses Sustainable Utilization (LMB-CAS), Guangdong Key Laboratory of Marine Materia Medica (LMMM-GD), South China Sea Institute of Oceanology, Chinese Academy of Sciences, Guangzhou 510301, People’s Republic of China.     

Arsenic detoxification potential of <Emphasis Type="Italic">aox</Emphasis> genes in arsenite-oxidizing bacteria isolated from natural and constructed wetlands in the Republic of Korea

Arsenic is subject to microbial interactions, which support a wide range of biogeochemical transformations of elements in natural environments such as wetlands. The arsenic detoxification potential of the bacterial strains was investigated with the arsenite oxidation gene, aox genotype, which were isolated from the natural and constructed wetlands. The isolates were able to grow in the presence of 10 mM of sodium arsenite (As(III) as NaAsO₂) and 1 mM of d+glucose. Phylogenetic analysis based on 16S rRNA gene sequencing indicated that these isolated strains resembled members of the genus that have arsenic-resistant systems (Acinetobacter sp., Aeromonas sp., Agrobacterium sp., Comamonas sp., Enterobacter sp., Pantoea sp., and Pseudomonas sp.) with sequence similarities of 81–98%. One bacterial isolate identified as Pseudomonas stutzeri strain GIST-BDan2 (EF429003) showed the activity of arsenite oxidation and existence of aoxB and aoxR gene, which could play an important role in arsenite oxidation to arsenate. This reaction may be considered as arsenic detoxification process. The results of a batch test showed that P. stutzeri GIST-BDan2 (EF429003) completely oxidized in 1 mM of As(III) to As(V) within 25–30 h. In this study, microbial activity was evaluated to provide a better understanding of arsenic biogeochemical cycle in both natural and constructed wetlands, where ecological niches for microorganisms could be different, with a specific focus on arsenic oxidation/reduction and detoxification.     

一株菲降解菌的特性及相关降解基因的克隆

从石油污染土壤中分离到一株菲降解菌2F5-2.根据该菌株生理生化特征和16S rDNA序列相似性分析,将其初步鉴定为鞘氨醇杆菌属(Sphingobium sp.).该菌株在10 h内对100 mg/L的菲的降解率为100%.降解菲的最适温度为30℃,最适pH为7.对降解途径的初步研究显示,该菌株通过水杨酸途径降解菲.克隆了编码芳香烃双加氧酶α亚基的基因phdA,它与菌株Sphingomonas sp.P2、Sphingobium yanoikuyae B1、Sphingomonas sp.ZP1中phdA的同源性分别为97.9%、98%和100%,表明该基因具有保守性.图6参16     

Isolation and characterization of a bacterial strain that efficiently degrades sex steroid hormones

A bacterial strain, ZY3, growing on sex steroid hormones as the sole source of carbon and energy was isolated from the sewage treatment plant of a prophylactic steroids factory. ZY3 degrades the 3-methoxy-17β-hydroxy-1,3,5(10),8(9)-δ-4-estren (MHE). This strain was preliminarily identified as Raoultella sp. ZY3 according to its morphology and its 16S rRNA gene sequence. During the experimental period (72 h), the optimum temperature, pH and 3-MHE concentration for the degradation of hydride by the strain ZY3 were 35°C, 10 and 10 mg/L, respectively. The degradation rate of the sex steroid hormones increased to 87% and 85% after the addition of maltose and peptone, respectively.     

Selection of poly(R)-3-hydroxybutyric acid utilising bacteria by enrichment,optimisation and compatibility testing for consortia development

Bacterial strains with poly-3-hydroxybutyrate (PHB) degradation potential were isolated from waste yard soil samples of selected industrial sites in Uttarakhand, India, and two microbial consortia were developed, i.e. Consortium I comprises Pseudomonas sp. strain Rb10, Pseudomonas sp. strain Rb11 and Bacillus sp. strain Rb18, and Consortium II is composed of Lysinibacillus sp. strain Rb1, Pseudomonas sp. strain Rb13 and Pseudomonas sp. strain Rb19. The current study involved enrichment selection via liquid and semi-solid media, followed by isolation and screening of bacterial strains using PHB pellets and films. Furthermore, the identification and characterisation was done by triphasic approach. The utilisation of PHB by the characterised strains was confirmed by Fourier transform infrared spectroscopy and scanning electron microscopy. Moreover, the minimum inhibitory concentration of solubilised PHB was found to be 2.5?mg/mL, which was detected through ‘clear zone assay’. Further, the selection and biocompatibility testing of potential isolates were performed for the formation of bacterial consortia. Thus, the present work would provide direct and standardised protocol for screening and selection of potential microbiomes for biodegradation of polymers by overcoming the negative effect of organic solvents. Moreover, indigenously developed consortia would be evaluated for their in situ biodegradation potential against various bioplastic films.     

Degradation of dibutyl phthalate in refuse from different phases of landfill by its dominant bacterial strain,Enterobacter sp. T1

A facultative bacterial strain isolated from municipal solid waste (MSW) obtained from a simulated landfill bioreactor was found to have the ability to use dibutyl phthalate (DBP) as its sole source of carbon and energy. Based on its morphology, physiochemical characteristics, and 16S rDNA sequence, the strain was identified as Enterobacter sp. T1. Evaluation of the degradation of DBP in refuse collected during the initial, acidic, and methanogenic phases of landfill before and after inoculation with Enterobacter sp. T1 revealed that the degradation fits first-order kinetic models for refuse from all phases. The removal rate of DBP in the refuse of the methanogenic phase increased from 59.3% to 74.5% when Enterobacter T1 was added. The half-life of DBP in refuse from the methanogenic phase that was inoculated with Enterobacter T1 decreased by 36.7% relative to uninoculated samples, and the intermediate products monobutyl phthalate (MBP) and phthalic acid were detected in all samples. These results provide new evidence for the potential of applying Enterobacter sp. for phthalic acid ester-polluted area remediation.     

Larval metamorphosis of the mussel <Emphasis Type="Italic">Mytilus galloprovincialis</Emphasis> in response to <Emphasis Type="Italic">Alteromonas</Emphasis> sp. 1: evidence for two chemical cues?

Bacterial isolates from multi-species biofilms were identified by 16S rDNA gene sequences and investigated for their inductive effects as monospecific biofilms on larval metamorphosis of Mytilus galloprovincialis. Alteromonas sp. 1 biofilm was found to have inductive activity, which increased with increasing cell density. The cue(s) of Alteromonas sp. 1 biofilm responsible for inducing larval metamorphosis was further investigated. Treatment of the biofilm with formalin, ethanol, heat or ultraviolet irradiation resulted in a significant reduction in the inductive activity of Alteromonas sp. 1, and the crude extract of surface-bound products of the biofilm showed no activity. These results indicated that if the cue was a surface-bound chemical cue, it was unstable, or susceptible to the treatments or the extraction process. On the other hand, the inductive activity of treated biofilms had a linear regression to the cell survival of bacteria, indicating a metabolically active biofilm was a requirement for larval metamorphosis. Conditioned water of the biofilm did not induce larvae to metamorphose. However, larval crawling behavior in the conditioned water was the same as that in the biofilm prior to larval metamorphosis, and significantly different to larval behavior in seawater. This indicated that a potential or partial waterborne cue existed, but remained inactive when alone. A synergistic effect of the conditioned water with formalin-fixed Alteromonas sp. 1 biofilm resulted in a significant increase in larval metamorphosis. Heat treatment and fractionation of the conditioned water demonstrated that the waterborne cue was heat-stable and <3,000 Da in molecular weight. Platinum-coating, Lentil Lectin and Wheat Germ Agglutinin treatments of the formalin-fixed biofilm significantly reduced its synergistic effect with the conditioned water, suggesting that a surface-bound cue was present on the biofilm and that the cue might be associated with the bacterial exopolysaccharide or glycoprotein. Evidence presented here suggests that two chemical cues derived from bacteria act synergistically on larval metamorphosis of Mytilus galloprovincialis.     

无苯酚培养基分离到的降酚菌多样性的分子分析

用3种不含苯酚的培养基(YPG、10%YPG和LB)从上海焦化厂废水处理系统(A2/O法)好氧池的悬浮污泥分离到24株降酚菌株.通过16SrDNAPCR扩增产物的限制性酶切分析(amplifiedribosomalDNArestrictionanalysis,ARDRA)、ERICPCR指纹图谱分析、多组分苯酚羟化酶大亚基(thelargestsubunitofthemulticomponentphenolhydroxylase,LmPH)基因的PCR扩增及16SrRNA基因测序的方法对这些降酚菌株进行表征.通过ARDRA分型,将这24株降酚菌株分为8个类型;利用ERICPCR可以将这24个菌株分为17种类型,说明同一ARDRA类型内菌株具有多样性.对17个ERIC类型代表菌株的16SrDNA扩增产物进行克隆并测序,测序结果在GenBank和RDP中进行比对.结果表明,与这17个代表菌株同源性最高的菌中,有6株是未见报道具有降酚功能的菌株.在这24株降酚菌中,有19株在苯酚含量为200mg/L的MP培养基中培养5d,苯酚降解率为20%左右,其余5株苯酚降解率达到100%,且均属于Rhodococcus属.本研究在降酚菌生物多样性上作了有益的探索.图2表1参20     

Modified dehydrogenase enzyme assay for evaluation of the influence of Hg,Cd, and Zn on the bacterial community structure of a wastewater treatment plant

In this study, a traditional assay was modified to evaluate the effect of Hg, Cd, and Zn on the bacterial community of a sequencing batch reactor and activated sludge plants and heavy metal-resistant bacterial species were determined. After the isolation of metal-resistant bacteria, their 16S rRNA gene fragments were sequenced. The BLAST program was used to compare the resulting 16S rRNA sequences with those in GenBank database to identify the isolated bacterial species. Hg was found to be the most toxic metal for both bacterial communities investigated. Sequence batch reactor bacteria were comparatively more resistant to Hg, Cd, and Zn than those from activated sludge. The resistant strains were close to the members of genus Pseudomonas, Kocuria, Stenotrophomonas, Enterococcus, and Staphylococcus. The modified dehydrogenase enzyme assay seems to be simple, robust, and competent for evaluation of the impact of metals on bacterial activity. Sequencing batch reactor systems should be preferred over activated sludge when wastewaters containing hazardous metals are to be treated.     

Bacterial communities of the marine sponges Hymeniacidon heliophila and Polymastia janeirensis and their environment in Rio de Janeiro, Brazil

In this study we performed a survey of the bacterial communities associated with the Western Atlantic demosponges Hymeniacidon heliophila and Polymastia janeirensis, based on 16S rRNA sequencing and transmission electron microscopy (TEM). We compared diversity and composition of the sponge-associated bacteria to those of environmental bacteria, represented by free-living bacterioplankton and by bacteria attached to organic particulate matter in superficial sediments. Partial bacterial 16S rRNA sequences from seawater, sediment, and sponges were retrieved by PCR, cloning, and sequencing. Sequences were subjected to rarefaction analyses, phylogenetic tree construction, and LIBSHUFF quantitative statistics to verify coverage and similarity between libraries. Community structure of the free-living bacterioplankton was phylogenetically different from that of the sponge-associated bacterial assemblages. On the other hand, some sediment-attached bacteria were also found in the sponge bacterial community, indicating that sponges may incorporate bacteria together with sediment particles. Rare and few prokaryotic morphotypes were found in TEM analyses of sponge mesohyl matrix of both species. Molecular data indicate that bacterial richness and diversity decreases from bacterioplankton, to particulate organic sediment, and to H. heliophila and P. janeirensis. Sponges from Rio de Janeiro harbor a pool of novel and exclusive sponge-associated bacterial taxa. Sponge-associated bacterial communities are composed of both taxons shared by many sponge groups and by species-specific bacteria.     

Strengthening effects of ammonia nitrogen on the harmless biological treatment of oily sludge

To improve the efficiency of oil degradation and strengthen the harmless treatment of oily sludge, three dominant strains identified as Chryseomicrobium sp. YL2, Gordonia sp. YL3 and Acinetobacter sp. YL5 were isolated from soil near a refinery, and the effects on the bioremediation of the oily sludge from the refinery were investigated. The results showed that the efficiency of oil degradation increased by 31.5% compared with the control when the dominant strains were added to the treatment of oily sludge. Furthermore, the dominant strains could use oil as a carbon source for heterotrophic nitrification–aerobic denitrification. The addition of ammonia nitrogen resulted in a large number of remaining microbes and heightened dehydrogenase activity in the oily sludge, further accelerating oil degradation, mainly for C₁₁ to C₂₅ saturated hydrocarbons, and the oil degradation efficiency increased by 40.8%. After 120 days of bioremediation, the biotoxicity of oily sludge, which was expressed by the equivalent phenol concentration, decreased by 40.0% compared with that of the control, indicating that the addition of ammonia nitrogen enhanced the biodegradation of oil. This method can be used to strengthen the harmless treatment of oily sludge in practical engineering applications.     

Isolation,identification, and the degradation characteristics of a thiobencarb-degrading bacterium Bacillus sp. T2北大核心CSCD

Thiobencarb, a thiocarbamate herbicide, is widely used to control weeds in rice paddies. Screening for highly efficient thiobencarb-degrading bacteria is important for the bioremediation of thiobencarb-contaminated environments. The aim of this study was to isolate and identify highly efficient thiobencarb-degrading bacteria and to identify the degradation pathway and the degrading properties. The thiobencarb-degrading strain was isolated using methods of microbiological acclimation and enrichment and was then identified using a 16S rRNA phylogenetic analysis. The degrading properties of the isolated bacterium were determined by single-factor experiments, and the degradation products were identified using gas chromatography-mass spectrometry (GC-MS). A thiobencarb-degrading strain T2, which can utilize thiobencarb as the sole source of carbon for energy and growth, was isolated from paddy soil. Strain T2 degraded more than 98.3% of 0.4 mmol/L of thiobencarb within 36 h. It was preliminarily identified as Bacillus sp. T2 according to the 16S rRNA gene analysis and from its morphological, physiological, and biochemical characteristics. The metabolic products of the thiobencarb degradation for strain T2 were identified as 4-chlorobenzyl mercaptan, 4-chlorobenzaldehyde, and 4-chlorobenzoic acid by the GC-MS. Based on metabolite identification, it was speculated that thiobencarb degradation in strain T2 was initiated by the hydrolysis of the thioester bond to produce 4-chlorobenzyl mercaptan, which was further oxidized to 4-chlorobenzaldehyde and 4-chlorobenzoic acid. The thiobencarb degradation that was initiated by the hydrolysis of the thioester bond by strain T2 is a new metabolic pathway, which provides valuable research material and reliable experimental data for revealing the metabolic process and mechanism of thiobencarb microbial degradation in soil. The strain Bacillus sp. T2 has a very high degradation efficiency, suggesting it is a good prospect for microbial remediation in thiobencarb-polluted environments. © 2018 Science Press. All rights reserved.     

Biodegradation of anthracene and phenanthrene by bacteria isolated from oil-contaminated soil of Bangladesh

Polycyclic aromatic hydrocarbons (PAHs) are ubiquitous environmental pollutants entering into the environment through natural and anthropogenic activities. Owing to their toxicity to various life forms including humans, detoxification of PAHs is crucial to reduce their effects on cells. In this study, we have isolated two bacteria capable of degrading two common PAHs, anthracene and phenanthrene, from contaminated soil samples by using selective enrichment culture supplemented with test PAHs as the sole source of carbon and energy. The isolated bacteria were identified and affiliated as Pseudomonas aeruginosa strain KD and Stenotrophomonas maltophilia strain RC based on their 16s rRNA gene sequences. The degradation of anthracene and phenanthrene was estimated indirectly by measuring the decolourisation extent of a redox indicator, 2, 6-dichlorophenolindophenol, incorporated into PAH-supplemented mineral salt media. In the case of anthracene, ≥90% decolourisation was recorded at 20 and 48 days for P. aeruginosa and S. Maltophilia, respectively. On the other hand, ≥94% decolourisation was recorded at 56 and 52 days for P. aeruginosa and S. maltophilia, respectively during the utilisation of phenanthrene.     

Biodegradation of phenylurea herbicide diuron by microorganisms from long-term-treated sugarcane-cultivated soils in Kenya

The phenylurea herbicide diuron [N-(3,4-dichlorophenyl)-N,N-dimethylurea] is widely used alone or in a broad range of herbicide formulations. Its degradation in sugarcane-cultivated soils which have been impacted by the herbicide through repeated applications was studied. Liquid culture experiments with diuron as the only carbon source led to the isolation of different bacterial strains capable of degrading diuron. The bacterial species belonging to the genera Bacillus, Vagococcus, and Burkholderia, identified through biochemical and molecular characterization, degraded diuron to different extents. The isolated Bacillus cereus, Vagococcus fluvialis, Burkholderia ambifaria, and Bacillus spp1 degraded diuron by 21%, 25%, 22%, and 19% of the initially applied concentration of 40?mg?L^?1, respectively, after 35 days of incubation in liquid culture media. Small amounts of 3,4-dichloroaniline and the de-methylated metabolite N-(3,4-dichlorophenyl)-N-methylurea were detected in liquid culture media. The combination of V. fluvialis and B. ambifaria showed an enhanced degradation of up to 30% of the initially applied concentration of 40?mg?L^?1. Degradation by pure isolates was low (18–25%) compared to the capacities of diuron degradation shown by the bacterial communities (58–74%). This study showed the presence of diuron degraders in sugarcane-cultivated soils impacted by diuron due to repeated applications.     

In situ enhanced bioremediation of dichlorvos by a phyllosphere Flavobacterium strain

A bacterium capable of degrading dichlorvos was isolated from the rape phyllosphere and designated YD4. The strain was identified as Flavobacterium sp., based on its phenotypic features and 16S rRNA gene sequence. Strain YD4 was able to utilize dichlorvos as the sole source of phosphorus. In situ enhanced bioremediation of dichlorvos by YD4 was hereafter studied. Chlorpyrifos and phoxim could also be degraded by this strain as the sole phosphorus source. A higher degradation rate of dichlorvos was observed after spraying YD4 onto the surface of rape leaves when compared to the sterilized-YD4 and water-treated samples. The results indicated that pesticide-degrading epiphytic bacterium could become a new way for in situ phyllosphere bioremediation where the hostile niche is unsuitable for other pesticide-degrading bacteria isolated from soil and water.     

假单胞菌AEBL3对土壤中呋喃丹的生物降解

从农药污染的农田土壤中分离到一株假单胞菌AEBL3，该菌能够以呋喃丹为唯一的碳源和氮源生长。使用AEBL3作为生物强化剂对模拟呋喃丹污染的土壤环境进行了生物修复试验。结果表明，接种AEBL3能够明显增加土壤中呋喃丹的降解率。降解菌AEBL3在土壤中具有一定的移动性，当从土壤表层加菌时，对0-7cm深土层中的呋喃丹都有很好的降解效果。在各种投加方式的比较中，以投加降解菌原液修复效果最好，缓冲液悬浮的菌细胞次之，砂粒混合物的效果最差。16S rRNA的变性梯度电泳(DGGE)结果揭示了生物修复过程中土壤微生物菌群结构中的动态变化。