Copper-based fungicide copper hydroxide accelerates the evolution of antibiotic resistance via gene mutations in Escherichia coli.

The extensive use of copper-based fungicides in orchards, especially in vineyards, leads to the accumulation of copper, which has caused growing concern. However, data on the acquisition of antibiotic resistance in opportunistic pathogens under copper-based fungicides are scarce. In this study, we investigated the potential development of antibiotic resistance in Escherichia coli K12 under selective copper hydroxide pressure. The results indicated that copper hydroxide at concentrations of 100 mg/L and 200 mg/L evolved resistance against chloramphenicol and tolerance against tetracycline to 4-8 and 2.00-2.67 times than the initial minimal inhibitory concentrations (MICs), respectively. Whole-genome sequencing analysis showed that the obtained resistant strains carried gene mutations including AcrAB-TolC multidrug efflux pump (acrB and marR), outer membrane porin (evZ), and another indirect pathways. Furthermore, the expression of multidrug efflux pump genes and oxidative stress-related genes were significantly upregulated, whereas outer membrane porin genes were downregulated. Thus, our results could well explain the emergence of antibiotic resistance and resistance mechanisms selected by copper-based fungicide, and provide a basis for the management of copper-based fungicide in agriculture to avoid the spread of antibiotic resistance.

Enantioselectivity of new chiral triazole fungicide mefentrifluconazole: Bioactivity against phytopathogen, and acute toxicity and bioaccumulation in earthworm (Eisenia fetida).

Elaborating the environmental behavior of mefentrifluconazole, a novel triazole fungicide, in stereoselective level is of paramount importance for the application of the pesticide in agriculture. In this study, the enantioselective bioactivity, acute toxicity and stereoselective bioaccumulation of mefentrifluconazole in earthworm (Eisenia fetida) were investigated. Bioactivity tests against four pathogens revealed that R-(-)-mefentrifluconazole exhibited approximately 11-113 times higher bioactivity than its S-(+)-mefentrifluconazole. However, the LC50 of S-(+)-, rac- and R-(-)-mefentrifluconazole to earthworm was measured to be 4.1, 11.4 and 7.3 µg/cm2, respectively, indicating active ingredient R-(-)-mefentrifluconazole is less toxic than its racemate and S-form. Accumulation of mefentrifluconazole in earthworms was non-enantioselective and negatively related to its adsorption onto soils. The concentration of mefentrifluconazole in in situ pore water (CIPW) and CaCl2 extraction (CCaCl2) was closely related to its accumulation in earthworms, suggesting that CIPW and CCaCl2 could be appropriate indicators for estimation of the bioavailability of mefentrifluconazole in soil. Conclusively, our study provides necessary information for the risk assessment of mefentrifluconazole in agriculture.

Emergence of Triazole Resistance in Aspergillus fumigatus Exposed to Paclobutrazol.

As a global health problem, the source of triazole resistance in Aspergillus fumigatus has gained much attention. This study was conducted to explore whether the triazole plant regulator paclobutrazol could evolve triazole resistance in A. fumigatus. The results indicated that two triazole-resistant strains with hereditary stability were isolated from liquid medium and soil. The up-regulation of cyp51A, cyp51B, AtrF, cdr1B, AfuMDR1, AfuMDR2, and AfuMDR4 played an important role in these resistant strains. The triazole-resistance in A. fumigatus could depend on the selective pressure of paclobutrazol concentration and exposure time. These results indicate that the application of paclobutrazol may result in the emergency of triazole resistance in A. fumigatus and thus have a potential risk for the treatment of invasive aspergillosis.

Prevalence of Azole-Resistant Aspergillus fumigatus is Highly Associated with Azole Fungicide Residues in the Fields.

Triazole resistance in Aspergillus fumigatus is a growing public health concern. In addition to its emergence in the therapy of invasive aspergillosis by triazole medicines, it has been frequently detected in agricultural fields all over the world. Here, we explore the potential link between residues of azole fungicides with similar chemical structure to triazole medicines in soil and the emergence of resistant A. fumigatus (RAF) through 855 500 km2 monitoring survey in Eastern China covering 6 provinces. In total, 67.3%, 15.2%, 12.3%, 2.9%, 1.5%, 0.4%, and 0.3% of the soil samples contained these five fungicides (tebuconazole, difenoconazole, propiconazole, hexaconazole, and prochloraz) of 0-100, 100-200, 200-400, 400-600, 600-800, 800-1000, and >1000 ng/g, respectively. The fractions of samples containing RAF isolates were 2.4%, 5.2%, 6.4%, 7.7%, 7.4%, 14.3%, and 20.0% of the samples with total azole fungicide residues of 0-100, 100-200, 200-400, 400-600, 600-800, 800-1000, and >1000 ng/g, respectively. We find that the prevalence of RAFs is positively (P < 0.0001) correlated with residual levels of azole fungicides in soils. Our results suggest that the use of azole fungicides in agriculture should be minimized and the intervals between treatments expanded to reduce the selective pressure toward the development of resistance in A. fumigatus in agricultural fields.

Chemical factors affecting uptake and translocation of six pesticides in soil by maize (Zea mays L.).

Uptake of residual pesticides in a soil by a certain crop plant may be governed by their physicochemical properties. Uptake and translocation of pesticides (imidacloprid, acetamiprid, tricyclazole, azoxystrobin, tebuconazole and difenoconazole) with the octanol/water partition coefficient (log Kow) ranging from 0.57 to 4.36 were investigated in soil with maize as a model plant. The results show that all tested pesticides in soil were uptaken by maize with accumulation amount of 27.73, 17.75, 18.96, 12.56, 10.66 and 2.13 µg for imidacloprid, acetamiprid, tricyclazole, azoxystrobin, tebuconazole and difenoconazole at 14 d, respectively. The accumulation amount was negatively correlated with adsorption coefficients and positively correlated with pesticide concentration in in situ pore water (CIPW). Root bioconcentration factor varied widely from 0.61 for imidacloprid to 974.64 for difenoconazole was positively correlated with log Kow and molecular weight but negatively with water solubility. Conversely, translocation factor varied from 0 for difenoconazole to 1.64 for imidacloprid was negatively correlated with log Kow but positively with water solubility. It determined that uptake, accumulation and translocation of the pesticides in soil by maize are governed by their physicochemical properties, especially log Kow. CIPW is an appropriate candidate to evaluate the accumulation of pesticides in maize from soil.

Characterization, genome functional analysis, and detoxification of atrazine by Arthrobacter sp. C2.

Residual injury of atrazine to the succeeding crops has been frequently reported. It is necessary to find a solution for the detoxification of atrazine contaminated soil. A high-efficient bacterial strain Arthrobacter sp. C2 for atrazine degradation was isolated in this study. The genomic information of the isolate C2, and its degradation characteristics and potential application in detoxification of atrazine contaminated soil were investigated. The results indicated that the isolate C2 genome contained 4,305,216 bp nucleotides, three plasmids, and 4705 coding genes. The degradation rates of atrazine at levels of 1, 10, 100 mg/L by the isolate C2 were 0.34, 1.94, 18.64 mg/L/d, respectively. The optimum temperature and pH for the isolate C2 to degrade atrazine were 30 °C and 7.0-9.0. Based on the metabolites detected by UPLC-TOF-MS/MS and genome annotation of the isolate C2, a common metabolic pathway of atrazine was proposed as that atrazine is firstly dechlorinated into hydroxyatrazine, and subsequently to N-isopropylammelide via dealkylation, and ultimately deaminated to cyanuric acid. Introduction of the isolate C2 into soil can enhance degradation of atrazine and thus eliminate the toxic effect of this herbicide on wheat growth. Our results indicate that the strain C2 could be a potential bioresource for bioremediation of atrazine contaminated soil.

Root Uptake of Imidacloprid and Propiconazole Is Affected by Root Composition and Soil Characteristics.

Residual pesticides in soil may be taken up by crops and negatively affect food safety. The uptake mechanism of imidacloprid and propiconazole was studied using wheat roots. The factors affecting root uptake were also studied with different crops and in different soils. Imidacloprid and propiconazole were taken up by wheat roots mainly through the symplastic and apoplastic pathways, respectively. Root protein and lipid contents were the main factors affecting the uptake and accumulation of imidacloprid and propiconazole by different crop roots, respectively. The uptake of imidacloprid and propiconazole in soil by wheat plants was linearly correlated with their concentrations in soil pore water, which were governed by soil characteristics. These results are helpful for understanding and estimating crop uptake of residual pesticides in soils.

Five-Year Survey (2014 to 2018) of Azole Resistance in Environmental Aspergillus fumigatus Isolates from China.

A total of 191 soil samples from Hangzhou, China, were submitted to detect non-wild-type (non-WT) Aspergillus fumigatus and its associated mechanisms. There were 2 (4.7%), 13 (12.4%), and 31 (23.1%) isolates identified as non-WT in 2014, 2016, and 2018, respectively. The resistant mutations of TR34/L98H, TR46/Y121F/T289A, and TR34/L98H/S297T/F495I were found in 3, 5, and 5 non-WT isolates. The G448S mutation, previously only found in clinical settings, was detected in A. fumigatus from soil samples.

Triazole resistance in Aspergillus fumigatus in crop plant soil after tebuconazole applications.

Aspergillus fumigatus is the primary agent of invasive aspergillosis (IA) causing high morbidity and mortality in immunocompromised patients. Triazole resistance in A. fumigatus and its sources have gained wide attention. For several years, environmental fungicides use has been proposed as the major cause for triazole resistance in A. fumigatus. However, there are few studies on azole-resistant A. fumigatus (ARAF) selected by triazole fungicides in agricultural systems. We studied the possible emergence of ARAF in the field after exposure to triazole fungicide tebuconazole. Our results showed that exposure to tebuconazole in soil selects for resistance to triazoles in A. fumigatus. The probability of ARAF developing in soils depends upon the concentrations of tebuconazole after application. We suggest that tebuconazole applications should be minimized to reduce selective pressure for the generation of ARAFs.

Increased triazole-resistance and cyp51A mutations in Aspergillus fumigatus after selection with a combination of the triazole fungicides difenoconazole and propiconazole.

Triazole-resistance in Aspergillus fumigatus is widespread. We evaluated whether triazole-resistance in A. fumigatus and its related cyp51A mutations, induced by a combination of the triazole fungicides difenoconazole and propiconazole, differs from resistance induced by the individual fungicides. Both difenoconazole and propiconazole can induce triazole-resistance in A. fumigatus. Resistance is much easier induced by formulated fungicides or a combination of these two fungicides compared with standard fungicides or individual fungicides, respectively. Six different mutations (G138S, G138D, H147Y, I246M, M263I and D430N) were identified in the induced resistant strains. The H147Y, I246M and M263I mutations were associated with triazole-resistance. This implies that the application of a combination of difenoconazole and propiconazole may result in higher triazole-resistance in A. fumigatus and more mutations in the cyp51A gene.

Inoculation of plant growth-promoting bacteria Bacillus sp. YM-1 alleviates the toxicity of Pb to pakchoi.

Heavy metal accumulation in plants may imperil human health. Inoculation of plant growth-promoting bacteria can alleviate the toxicity of heavy metal and promote plant growth. In this study, Bacillus sp. YM-1, a heavy metal resistant and plant growth-promoting bacterium, was immobilized with spent substrate of mushroom and applied to alleviate the toxicity of Pb to pakchoi. The results indicated that the biomass of pakchoi in inoculation group was increased by 17.45 to 27.05% compared with that in non-inoculated group (p < 0.05). The root and shoot were lengthened by 13.45% to 39.17% and 20.23% to 42.36%, respectively. The content of Pb in root and shoot obviously reduced and that in edible part (shoot) was less than 0.2 mg kg-1 in the low concentration of Pb. Other indicators such as superoxide dismutase (SOD), peroxidase (POD), chlorophyll, and protein all testified that YM-1 inoculation was conducive to the alleviation of Pb toxicity to pakchoi.

Batch and fixed-bed biosorption of Cd(II) from aqueous solution using immobilized Pleurotus ostreatus spent substrate.

To prevent the blockage in a continuous fix-bed system, Pleurotus Ostreatus spent substrate (POSS), a composite agricultural waste, was immobilized into granular adsorbents (IPOSS) with polymeric matrix, and used to remove Cd(II) from synthetic wastewater in batch experiment as well as in continuous fixed-bed column system. In batch experiment, higher pH, temperature and Cd(II) initial concentration were conducive to a higher biosorption capacity, and the maximum biosorption capacity reached up to 87.2 mg/g at Cd(II) initial concentration of 200 mg/L, pH 6 and 25 °C. The biosorption of Cd(II) onto IPOSS followed the Langmuir isotherm model with the maximum adsorption capacity(qmax) of 100 mg/g. The biosorption was an endothermic reaction and a spontaneous process based on positive value of ΔH0 and negative value of ΔG0. In fixed-bed column system, higher bed depth, lower flow rate and influent Cd(II) concentration led to a longer breakthrough and exhaustion time, and the best performance (equilibrium uptake (qe) of 14.4 mg, breakthrough time at 31 h and exhaustion time at 78 h) was achieved at a bed depth of 110 cm, a flow rate of 1.2 L/h and an influent concentration of 100 mg/L. Furthermore, regeneration experiment revealed a good reusability of IPOSS with 0.1 M HNO3 as eluting agent during three cycles of adsorption and desorption. Cd(II) biosorption onto IPOSS mainly relied on a chemical process including ion exchange and complexation or coordination revealed by SEM-EDX, FTIR and XRD analysis.

Characterization of siderophore produced by Pseudomonas syringae BAF.1 and its inhibitory effects on spore germination and mycelium morphology of Fusarium oxysporum.

In this study, an antagonistic bacterium against Fusarium oxysporum was identified and designated as Pseudomonas syringae strain BAF.1 on the basis of 16S rDNA sequence analysis and physiological-biochemical characteristics. It produced catechol-species siderophore at a molecular weight of 488.59 Da and a maximum amount of 55.27 µg/ml with glucose as a carbon source and asparagine as a nitrogen source at a C/N ratio of 10:1, 30°C and pH 7. The siderophore exhibited prominent antagonistic activity against Fusarium oxysporum with a maximum inhibition rate of 95.24% and had also suppressive effects on other kinds of 11 phytopathogenic fungi in the absence of FeCl3·6H2O. Spore germination was completely inhibited by 50 µl of the siderophorecontaining solution, and the ultrastructures of mycelia and spores were also considerably suppressed by siderophore treatment as established by electron microscopy observation. These results indicate that the siderophore produced by Pseudomonas syringae BAF.1 could be potentially used for biocontrol of pathogenic Fusarium oxysporum.

Biosorption characteristic of Alcaligenes sp. BAPb.1 for removal of lead(II) from aqueous solution.

In this study, strain BAPb.1 was isolated from lead mining area and used as an adsorbent to remove lead(II) ions from aqueous solution. The physicochemical characteristics, heavy metal resistance and antibiotic sensitivity of strain BAPb.1 were investigated. Biosorption capacity was evaluated by batch biosorption experiments, and isothermal characteristics were discussed. Atomic force microscopy (AFM), scanning electron microscopy (SEM), energy dispersive X-ray spectroscopy (EDX) and Fourier transform infrared spectrometry (FTIR) were conducted to explore the mechanism for lead(II) adsorption. Based on morphological and physiological characteristics as well as the phylogenetic analysis of 16S rDNA sequences, strain BAPb.1 was identified as a member of the genus Alcaligenes. It exhibited high resistances to multiple heavy metals such as lead(II), copper(II), zinc(II), nickel(II) and chromium(VI), and to antibiotics such as kanamycin, ampicillin, streptomycin, chloramphenicol, and tetracycline. The optimum conditions for maximum biosorption rate of 85.2% and maximum capacity of 56.8 mg g-1 were found at pH of 5, adsorbent dosage of 1.5 g L-1 (dry weight), initial lead(II) concentration of 100 mg L-1, and contact time of 30 min at 30 °C. Biosorption isotherms were well fitted with Langmuir isotherm model. Mechanism analysis reveals that the lead(II) ions may exchange with sodium and potassium ions, and the hydroxyl, carbonyl and phosphate groups on the cell surface can chelate the lead(II) ions, therefore, surface adsorption play significant role in the biosorption process.

Optimization of Siderophore Production by Bacillus sp. PZ-1 and Its Potential Enhancement of Phytoextration of Pb from Soil.

In this study, the siderophore-producing characteristics and conditions of Bacillus sp. PZ-1 were investigated and the enhancement of siderophores on Pb uptake and translocation in Brassica juncea were determined. Results of single factor experiment showed that glucose, pH, and Pb(NO3)2 could stimulate PZ-1 growth and siderophore production. The maximum siderophore production of 90.52% siderophore units was obtained by response surface methodology optimization at the glucose concentration of 21.84 g/l, pH 6.18, and Pb(NO3)2 concentration of 245.04 µmol/l. The type of siderophore was hydroxamate and its concentration in the fermentation broth amounted to 32.24 µg/ml. Results of pot experiments indicated that the siderophores enhanced B. juncea to assimilate more Pb from soil with the uptake ratio from 1.04 to 2.74, and to translocate more Pb from underground to overground with the TF values from 1.21 to 1.48. The results revealed that Bacillus sp. PZ-1 could produce abundant siderophores and might be potentially used to augment the phytoextraction of Pb from soil.

Removal of Cr(VI) by surfactant modified Auricularia auricula spent substrate: biosorption condition and mechanism.

Auricularia auricula spent substrate (AASS) modified by didodecyldimethylammonium bromide(DDAB) was used as adsorbent to remove Cr(VI) from aqueous solution. Based on a single-factor experiment and response surface methodology, the optimal conditions were adsorbent dosage of 1.5 g/L, pH value of 4.0, initial Cr(VI) concentration of 19 mg/L, temperature of 25 °C, biosorption time of 120 min, rotational speed of 150 r/min, respectively, under which biosorption capacity could reach 12.16 mg/g compared with unmodified AASS (6.058 mg/g). DDAB modification could enlarge the specific surface area and porous diameter of the adsorbents, and supply hydrophilic and hydrophobic groups capable of adsorbing at the interfaces. In addition, DDAB increased ionic exchange and complex formation demonstrated by variations of elemental contents, shifts of carboxyl, amine groups, hydroxyl, alkyl chains, and phosphate groups as well as the crystal structure of the Cr-O compounds. Variations of peaks and energy in XPS analysis also testified the reduction of Cr(VI) to Cr(III).The biosorption behavior of modified AASS was in line with Langmuir and Freundlich isotherm equation. The final regeneration efficiency was 62.33% after three biosorption-desorption cycles. Apparently, DDBA is a eximious modifier and DDBA-modified AASS was very efficient for Cr(VI) removal.

Inonotus obliquus polysaccharide regulates gut microbiota of chronic pancreatitis in mice.

Polysaccharide is efficient in attenuation of metabolic ailments and modulation of gut microbiota as prebiotics. The therapeutic effect of Inonotus obliquus polysaccharide (IOP) on chronic pancreatitis (CP) in mice has been validated in our previous study. However, it is not clear whether IOP is conducive to maintaining the homeostasis between gut microbiota and host. The aim of this study is to testify the potential effects of IOP on gut microbiota composition and diversity in mice with CP. The changes in glutathione peroxidase (GSH-PX), total antioxidant capacity (TAOC), tumor necrosis factor alpha (TNF-α), transforming growth factor beta (TGF-ß), lipase and trypsin levels were measured by commercial assay kits, meanwhile the gut microbiota composition and diversity were analyzed by high throughput sequencing. The IOP treatment increased GSH-PX and TAOC levels, and decreased TNF-α, TGF-ß, lipase and trypsin levels in CP mice. It was also observed that gut microbiota in IOP treated groups were less diverse than others in terms of lower Shannon diversity index and Chao 1 estimator. IOP increased the proportion of Bacteroidetes and decreased that of Firmicutes at phylum level. Bacteroidetes was found positively correlated with GSH-PX and TAOC, and Firmicutes correlated with TNF-α, TGF-ß, and lipase. In conclusion, administration of IOP could regulate gut microbiota composition and diversity to a healthy profile in mice with CP, and some bacterial phylum significantly correlated with characteristic parameters.

Effects of Ganoderma lucidum polysaccharides on chronic pancreatitis and intestinal microbiota in mice.

This study manifested the effects of polysaccharides from Ganoderma lucidum strain S3 (GLP S3) on chronic pancreatitis (CP) therapy and intestinal microbiota modulation in mice induced by diethyldithiocarbamate (DDC). The GLPS3 was prepared from cultured mycelium and markedly alleviated the pancreatitis in mice through decreasing lipase, AMS, IFN-γ and TNF-α level as well as increasing SOD and total antioxidant activity. Furthermore, high throughput sequencing analysis revealed that GLPS3 altered the composition and diversity of intestinal microbiota, especially, decreased the relative abundance of phylum Bacteroidetes and increased that of phylum Firmictutes. At the genus level, supplementation of GLPS3 increased the relative abundance of the beneficial bacteria such as Lactobacillales, Roseburia and Lachnospiraceae. These results disclosed the potential therapy mechanism of GLPS3 on chronic pancreatitis might be intestinal microbiota dependent.