The formation kinetics and control of biofilms by three dominant fungi species isolated from groundwater.

Filamentous fungi can enter drinking water supply systems in various ways, and exist in suspended or sessile states which threatens the health of individuals by posing a high risk of invasive infections. In this study, the biofilms formation kinetics of the three genera of fungal spores, Aspergillus niger (A. niger), Penicillium polonicum (P. polonicum) and Trichoderma harzianum (T. harzianum) isolated from the groundwater were reported, as well as the effects of water quality parameters were evaluated. In addition, the efficiency of low- concentrations of chlorine-based disinfectants (chlorine, chlorine dioxide and chloramine) on controlling the formation of fungal biofilms was assessed. The results showed that the biofilms formation of the three genera of fungi could be divided into the following four phases: induction, exponential, stationary and sloughing off. The optimum conditions for fungal biofilms formation were found to be neutral or weakly acidic at 28 °C with rich nutrition. In fact, A. niger, P. polonicum, and T. harzianum were not observed to form mature biofilms in actual groundwater within 120 hr. Carbon was found to have the maximum effect on the fungal biofilms formation in actual groundwater, followed by nitrogen and phosphorus. The resistance of fungal species to disinfectants during the formation of biofilms decreased in the order: A. niger > T. harzianum > P. polonicum. Chlorine dioxide was observed to control the biofilms formation with maximum efficiency, followed by chlorine and chloramine. Consequently, the results of this study will provide a beneficial understanding for the formation and control of fungal biofilms.

MiR-9-5p inhibits mitochondrial damage and oxidative stress in AD cell models by targeting GSK-3ß.

This study aims to investigate the effects and underlying mechanisms of overexpression microRNA-9-5p (miR-9-5p) on the Aß-induced mouse hippocampal neuron cell line HT22. Different concentrations of Aß25-35 (10, 20, 40, 80, and 160 µM) treatment were used to establish AD model in HT22 cells. The CCK-8 assay was used to measure the cell viability. The mRNA expression levels of miR-9-5p and glycogen synthase kinase-3ß (GSK-3ß) were determined by RT-qPCR. HT22 cell apoptosis was analyzed flow cytometry. MiR-9-5p was down-regulated in Aß25-35-induced HT22 cells. GSK-3ß is a functional target for miR-9-5p. MiR-9-5p overexpression inhibited Aß25-35-induced mitochondrial dysfunction, cell apoptosis, and oxidative stress by regulating GSK-3ß expression in HT22 cells. Furthermore, through targeting GSK-3ß, overexpression of miR-9-5p partly activated nuclear factor Nrf2/Keap1 signaling, including part increases of Nrf2, HO-1, SOD-1, GCLC expression and slight decrease of Keap1 expression. Our results showed miR-9-5p may play a powerful role in the pathogenesis of AD.

Influence of heavy metal ions on the spectra and charge characteristics of DOM of municipal sewage secondary effluent.

The spectral and charged characteristics of dissolved organic matter (DOM) in municipal sewage secondary effluent influenced by heavy metal ions were studied by three-dimensional fluorescence spectroscopy, ultraviolet spectrometry, etc. As the Cu(II) concentration increased, the fluorescence intensity of the DOM's fluorescence peak reduced. When the Cu(II) concentration was 100 µmol/L, the fluorescence intensity of the humic-like acid in the DOM reduced by 19.5%-27.6%. However, there was no obvious change in the fluorescence intensity when Cd(II) was added. According to the modified Stern-Volmer equation, the conditional stability constant lgK and the proportion of the fluorescent functional groups of the DOM binding with Cu(II) were 2.19-2.69 and 16%-48%, respectively. The UV253/UV203 value changed little with the additional dosage of metal ions below 10 µmol/L, which increased as the concentration of Cu(II) or Pb(II) increased from 10-100 µmol/L. As the metal concentrations increase, the absolute values of the Zeta potential and the particle sizes of the DOM basically increased. The results showed that Cu(II) had the strongest binding ability to the groups that emitted fluorescence in the secondary effluent DOM compared to Pb(II) and Cd(II).

A Micro-Scale Investigation on the Behaviors of Asphalt Mixtures under Freeze-Thaw Cycles Using Entropy Theory and a Computerized Tomography Scanning Technique.

The thermodynamic behavior of asphalt mixtures is critical to the engineers since it directly relates to the damage in asphalt mixtures. However, most of the current research of the freeze-thaw damage of asphalt mixtures is focused on the bulk body from the macroscale and lacks a fundamental understanding of the thermodynamic behaviors of asphalt mixtures from the microscale perspective. In this paper, to identify the important thermodynamic behaviors of asphalt mixtures under freeze-thaw loading cycle, the information entropy theory, an X-ray computerized tomography (CT) scanner and digital image processing technology are employed. The voids, the average size of the voids, the connected porosity, and the void number are extracted according to the scanned images. Based on the experiments and the CT scanned images, the information entropy evolution of the asphalt mixtures under different freeze-thaw cycles is calculated and the relationship between the change of information entropy and the pore structure characteristics is established. Then, the influences of different freezing and thawing conditions on the thermodynamic behaviors of asphalt mixtures are compared. The combination of information entropy theory and CT scanning technique proposed in this paper provides an innovative approach to investigate the thermodynamics behaviors of asphalt mixtures and a new way to analyze the freeze-thaw damage in asphalt mixtures.

Changes of characteristics and disinfection by-products formation potential of intracellular organic matter with different molecular weight in metalimnetic oxygen minimum.

Metalimnetic oxygen minimum (MOM) occurs in reservoirs or lakes due to stratification and algal blooms, which has low dissolved oxygen (DO) levels and leads to the deterioration of water quality. The transformation mechanism and the impact on the water quality of intracellular organic matter (IOM) derived from algae are poorly understood under MOM conditions. In this study, IOM extracted by Microcystis aeruginosa was divided into five components according to molecular weight (MW), and the changes of characteristics and correlated disinfection by-products formation potential (DBPFP) were analyzed and compared under MOM conditions. The removal efficiency of dissolved organic carbon (DOC) in the <5 kDa fraction (66.6%) was higher than that in the >100 kDa fraction (41.8%) after a 14-day incubation under MOM conditions. The same tendency also occurred in Fmax and DBPFP. The decrease in Fmax was mainly due to the decline in tryptophan-like and tyrosine-like for all IOM fractions. The diversity of microorganisms degrading the MW > 100 kDa fraction was lower than others. Besides low MW fractions, these findings indicated that more attention should be paid to high MW fractions which were resistant to biodegradation under MOM conditions during water treatment.

The aggregation characteristics of Aspergillus spores under various conditions and the impact on LPUV inactivation: Comparisons with chlorine-based disinfection.

Aggregation is the primary step prior to fungal biofilm development. Understanding the attributes of aggregation is of great significance to better control the emergence of waterborne fungi. In this study, the aggregation of Aspergills spores (A. flavus and A. fumigatus) under various salt, culture medium, and humic acid (HA) conditions was investigated for the first time, and the inactivation via low-pressure ultraviolet (LPUV) upon aggregated Aspergillus spores was also presented. The aggregation efficiency and size of aggregates increased over time and at low salt (NaCl and CaCl2) concentration (10 mM) while decreasing with the continuous increase of salt concentration (100 and 200 mM). Increasing the concentration of culture medium and HA promoted the aggregation of fungal spores. Spores became hydrated, swelled, and secreted more viscous substances during the growth period, which accelerated the aggregation process. Results also suggested that fungal spores aggregated more easily in actual water, posing a high risk of biohazard in real-life scenarios. Inactivation efficiency by LPUV decreased with higher aggregation degrees due to the protection from the damaged spores on the outer layer and the shielding of pigments in the cell wall. Compared to chlorine-based disinfection, the aggregation resulted in the extension of shoulder length yet neglectable change of inactivation rate constant under LPUV treatment. Further investigation of cell membrane integrity and intracellular reactive oxygen species was conducted to elucidate the difference in mechanisms between various techniques. This study provides insight into the understanding and controlling of the aggregation of fungal spores.

Occurrence and removal efficiency of six polycyclic aromatic hydrocarbons in different wastewater treatment plants.

Occurrence and removal efficiency of six polycyclic aromatic hydrocarbons (PAHs) (fluoranthene, benzo[b]fluoranthene, benzo[k]fluoranthene, benzo[a]pyrene, benzo[ghi]perylene and indeno[1,2,3-cd]pyrene) in wastewater were monitored at different treatment processes in three wastewater treatment plants (WWTPs) by using ultra performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). Chromatographic separation was performed on a Waters Acquity UPLC BEHC18 column (1.7 µm, 2.1 mm × 50 mm). A 0.2 µm precolumn filter was used to protect the analytical column. The result indicated that (1) the total concentrations of six PAHs were 427, 318, and 220 ng L(-1) in the influent of three wastewater treatment plants, respectively. Among six PAHs, the content of fluoranthene was the highest, accounting for 62-66% of total PAHs. (2) Throughout the wastewater treatment process, different treatment units exhibited different removal efficiencies for six PAHs. Biological treatment stage and aerated grit chamber appeared to be necessary for effective removal of six PAHs. The removal efficiencies of six PAHs were 73-83% at biological treatment units, and 24-56% at the aerated grit stage, respectively. The final sedimentation and disinfection for PAH removal was minor.

Investigation of the Relationship between Permanent Deformation and Dynamic Modulus Performance for Bearing-Layer Asphalt Mixture.

Of major concern is the lack of correlation between the material design and structural function of asphalt pavement in China. The objective of this paper is to identify the layer in asphalt pavement where permanent deformation occurs most seriously and to propose a control index for that layer's asphalt mixture. The permanent deformation of each layer was determined through the utilization of thickness measurements obtained from field cores. The results indicate that the reduction in thickness is more significant in the driving lane than in the ridge band and shoulder. This phenomenon can be attributed to the intensified densification and shearing deformation that arise from the combined impacts of recurrent axle loads and high temperatures. Compared to surface and base layers, the bearing layer is the primary area of concern for permanent deformation in asphalt pavement. Therefore, it is imperative to incorporate the ability of bearing-layer asphalt mixture to withstand permanent deformation as a crucial design parameter. The dynamic modulus of the bearing-layer asphalt mixture is significantly influenced by the type of asphalt, gradation, and asphalt content, compared to other design parameters. Based on the relationship established between dynamic modulus and dynamic stability, with creep rate as the intermediate term, a control standard was proposed to evaluate the permanent deformation of the bearing-layer asphalt mixture. This study can provide reasonable and effective guidance for prolonging pavement life and improving pavement performance.

The change of amino acids samples under metalimnetic oxygen minimum condition: Characterization and mechanism.

The metalimnetic oxygen minimum (MOM) is a common anaerobic phenomenon that occur between 5.00 and 40.00 m of reservoirs. Amino acids (AAs) are widely found in water, but their change in MOM remain unclear. In this study, four AAs with different side chain groups were selected to explore the change of their samples and related disinfection by-products formation potential (DBPFPs) under MOM condition. The results showed that the final degradation rate of dissolved organic carbon and dissolved organic nitrogen of four AAs samples were 11.71%-59.87% and 26.50%-100.00% under MOM condition. Aspartic acid samples were the easiest to be degraded, whereas glycine samples were the opposite. While the total fluorescence intensity increased by 6.30%-113.40% for the appearance of tryptophan-like substance. The total DBPFPs of glutamic acid, arginine and aspartic acid samples were finally decreased by 4.73%, 8.00% and 98.88% (glycine sample increased by 2.30 times). Compared with the surface condition, the degradation of AAs samples and the change of DBPFPs were significantly inhibited under MOM condition. In addition, the diversities of bacterial communities were significantly reduced under MOM condition, which was very unfavorable to the degradation of AAs samples, and in turn affected the control of DBPs and deteriorated the water quality.

Nondestructive Detection and Early Warning of Pavement Surface Icing Based on Meteorological Information.

Monitoring and warning of ice on pavement surfaces are effective means to improve traffic safety in winter. In this study, a high-precision piezoelectric sensor was developed to monitor pavement surface conditions. The effects of the pavement surface temperature, water depth, and wind speed on pavement icing time were investigated. Then, on the basis of these effects, an early warning model of pavement icing was proposed using an artificial neural network. The results showed that the sensor could detect ice or water on the pavement surface. The measurement accuracy and reliability of the sensor were verified under long-term vehicle load, temperature load, and harsh natural environment using test data. Moreover, pavement temperature, water depth, and wind speed had a significant nonlinear effect on the pavement icing time. The effect of the pavement surface temperature on icing conditions was maximal, followed by the effect of the water depth. The effect of the wind speed was moderate. The model with a learning rate of 0.7 and five hidden units had the best prediction effect on pavement icing. The prediction accuracy of the early warning model exceeded 90%, permitting nondestructive and rapid detection of pavement icing based on meteorological information.

Effective inactivation of fungal spores by the combined UV/PAA: Synergistic effect and mechanisms.

Peracetic acid (PAA) can effectively inactivate fungi in water, while may pose a potential risk of regrowth after disinfection. The inactivation kinetic and mechanism of fungal spores by combined UV and PAA (UV/PAA) was investigated in this study. The results showed that synergistic factor of the inactivation of A. niger and A. flavus was 1.44 and 1.37, which indicated significant synergistic effect of UV/PAA. The k of A. niger and A. flavus was similar at pH 5.0 and 7.0, while decreased 60.00% and 39.13% at pH 9.0 compared with that at pH 7.0. The effect of HA concentration on the inactivation efficiency of fungal spores by UV/PAA was negative, while the effect of PAA concentration was positive. The membrane permeabilized cell of A. niger and A. flavus caused by UV/PAA was 17.0% and 31.7%, which was higher than that caused by PAA and UV alone. The changes of morphology of fungal spores and the leakage of intracellular material indicated that the damage of cell structure caused by UV/PAA system was more serious than that of UV or PAA alone. In addition, the four parts that contributed in UV/PAA system was in the following order: UV > radical > PAA > synergistic effect. The inactivation efficiency of combined UV and chlorine (UV/Cl2) was higher than that of UV/PAA. Furthermore, the typical order of the inactivation efficiency in different matrix was: phosphate buffer solution > surface water > secondary effluent. The regrowth potential of fungal spores after UV/PAA treatment was significantly lower than that by PAA alone, indicating that UV/PAA could decrease the microbial regrowth potential after PAA disinfection alone.

Mutual-activation between Zero-Valent iron and graphitic carbon for Cr(VI) Removal: Mechanism and inhibition of inherent Side-reaction.

The low reactivity of zero-valent iron (ZVI) usually limits its application for pollutant remediation. Therefore, a microscopic galvanic cell (mGC) with short-circuited cathode and anode was synthesized to intensify its galvanic corrosion. The prepared mGC exhibited 7.14 times higher Fe(II) release performance than ordinary nanoscale-ZVI (nZVI), rendering efficient Cr(VI) removal performance. Density functional theory (DFT) revealed mutual-activation of the cathode and anode due to close proximity, dramatically enhancing the galvanic corrosion of Fe(0) in mGC. The corrosion potential of mGC was measured as -0.77 V, which was 100 mV more negative than nZVI. The released electrons and surface-bond Fe(II) from anode in mGC was proved to be the dominant reductive species. More importantly, Cr(VI) reduction was slightly inhibited by hydroxyl radicals generated by a series of inherent side-reactions in the system, which could be well eliminated by low concentrations of 4-acetamido phenol. This study provides a promising strategy for ZVI activation, and sheds light on its environmental applications.

Change of algal organic matter under different dissolved oxygen and pressure conditions and its related disinfection by-products formation potential in metalimnetic oxygen minimum.

Most of the reservoirs or lakes will form a metalimnetic oxygen minimum (MOM) with the characterization of a substantial fraction of dissolved oxygen (DO) depleted below the epilimnion. The effect of intracellular organic matter (IOM) of algal cells transformed under MOM conditions is completely different from that of the original IOM on water quality. In this study, the IOM changes of Microcystic aeruginosa under different MOM conditions and its related disinfection by-products formation potentials (DBPFPs) were investigated by changing the pressure and DO concentration of MOM. Total Fmax increased slightly and then decreased under different pressure conditions, finally decreasing by no more than 22.0%. Under aerobic condition, dissolved organic carbon (DOC) and total Fmax decreased significantly, and decreased by 60.4% and 38.8% within the first 2 days. The results of specific UV absorbance (SUVA) and UV250/UV365 indicated that aromatic compounds and average molecular weight of IOM were gradually increased under different MOM conditions. The total DBPFPs increased firstly and then decreased under different pressure conditions, and finally decreased by 26.2%-33.1%. The decrease of total DBPFPs was significantly higher under aerobic condition than that under anoxic condition, which finally decreased by 64.5%. Redundancy analysis showed that the fluorescence parameter (protein-like and humic-like fluorescence) could be expected as an index to predict the DBPFPs. Moreover, the results revealed that with the decrease of DO, the activity and diversity of natural microbial consortium decreased, which prevented the further degradation and utilization of organic matter by natural microbial consortium. Therefore, lower DO was a key player for the deterioration of water quality under MOM conditions.

Adding CuCo2O4-GO to inhibit bromate formation and enhance sulfamethoxazole degradation during the ozone/peroxymonosulfate process: Efficiency and mechanism.

In this work, a new type of catalyst CuCo2O4-GO was synthesized as a heterogeneous catalyst, and its control effect on bromate (BrO3-)generation and sulfamethoxazole (SMX) degradation in O3/PMS process was studied. When 100 mg/L CuCo2O4-GO was added to the reaction system, the BrO3- concentration generated was 0.25 µM at pH = 7.0, 100 µM PMS addition and 1.30 mg/min ozone injection after 30 min reaction. Compared with the 6.58 µM BrO3- produced in the control group, the addition of CuCo2O4-GO prominently inhibited the generation of BrO3- and the inhibition efficiency reached 96.17 %. The addition of CuCo2O4-GO inhibited the conversion of hypobromous acid, thereby inhibiting the formation of BrO3-. Meanwhile, the first-order kinetic constant of the degradation of SMX by O3/PMS and O3/PMS/CuCo2O4-GO was 0.163 and 0.422 min-1, respectively. The addition of CuCo2O4-GO promoted the degradation of SMX and the removal efficiency was reached above 98 % after 10 min reaction. According to the optimization of the GO loading ratio, it was found that CuCo2O4-GO with 20 % GO loading had the best promotion effect on the degradation of SMX, and almost completely inhibited the formation of BrO3-. Finally, in the repeated cycle experiment, CuCo2O4-GO could maintain its high catalytic activity and still had a high removal effect on SMX after three repeated uses. Besides, the BrO3- inhibition efficiency was above 80 % after two repeated uses. Therefore, adding synthetic CuCo2O4-GO is an effective way to control the formation of BrO3- and enhance the degradation of SMX in the O3/PMS process.

Efficacy of UV-LED based advanced disinfection processes in the inactivation of waterborne fungal spores: Kinetics, photoreactivation, mechanism and energy requirements.

The contamination of fungi in water supply systems poses great risks to environment and human health. In this work, UV light-emitting diodes (UV-LEDs)-based advanced disinfection processes (ADPs) including UV-LEDs/hydrogen peroxide (H2O2), UV-LEDs/persulfate (PS) and UV-LEDs/peroxymonosulfate (PMS), were adopted for waterborne fungal spores inactivation. Overall comparisons of the UV-LEDs-based ADPs with respect to the control efficiency of photoreactivation and energy consumption were also evaluated. Results showed that culturability reduction of the fungal spores treated by UV-LEDs was not enhanced with the addition H2O2, PMS, and PS according to the results of heterotrophic plate counts and reaction rate constants; A. niger was expected to have higher UV resistance followed by T. harzianum and P. polonicum. However, UV-LEDs-ADPs inactivation, especially at the wavelengths of 280 and 265/280 nm, could accelerate the permeabilization of fungal spores as characterized by flow cytometry. Take P. polonicum for example, the percentage of membrane permeabilized spores was 98.0%, 98.7%, 97.6% and 82.6% after treatment by UV280/H2O2, UV280/PS, UV280/PMS and UV280 alone, respectively at the fluence of 100 mJ/cm2. The direct attack of free radicals in the processes of UV-LEDs-ADPs further enhanced the membrane damage and lowered the photoreactivation level, thus improved the inactivation efficiency. UV-LEDs/H2O2 was considered as an effective process in the disinfection of fungal spores with the advantages of enhancing the damage of membrane, inhibiting photoreactivation and comparable energy consumption compared with UV-LEDs alone.

High performance liquid chromatography-tandem mass spectrometry for rapid and sensitive analysis of six polycyclic aromatic hydrocarbons in wastewater.

A sensitive and fast method was developed to quantitatively analyse the six polycyclic aromatic hydrocarbons (fluoranthene (FLT), benzo[b]fluoranthene (BbF), benzo[k]fluoranthene (BkF), benzo[a] pyrene (Bap), benzo[ghi]perylene (BghiP) and indeno[1,2,3-cd]pyrene (INPY)) by high performance liquid chromatography (UPLC) coupling with tandem mass spectrometry (MS/MS). Chromatographic separation was performed on a Waters Acquity UPLC BEHC18 column (1.7 microm, 2.1 mm x 50 mm). A 0.2 microm precolumn filter was used to protect the analytical column. Mobile phase Awas acetonitrile containing 0.5% toluene. Mobile phase B was water. Linearity of detection was in the range of 1-100 microg L(-1); LOD of 5 PAHs were lower than 0.1 microg L(-1); LOQ were 0.2 microg L(-1) except for benzo[k]fluoranthene. The LOD and the LOQ of benzo[k]fluoranthene were respectively 0.1 microg L(-1) and 0.8 microg L(-1). Wastewater samples collected from two wastewater treatment plants were determined using this method respectively. Recovery of all compounds varied from 67.8 +/- 10.6% to 113.2 +/- 7.2%. In comparison with the existing methods, this rapid method saves time and solvent and improves instrument sample throughput by 2-5 fold.

Synergistic effect of ozone and chlorine on inactivating fungal spores: Influencing factors and mechanisms.

Effective control of fungal contamination in water is vital to provide healthy and safe drinking water for human beings. Although ozone was highly effective in inactivating fungi in water, it was limited by a lack of continuous disinfection ability in water supply system. In present study, the inactivation of fungal spores by combining ozone and chlorine was investigated. The synergistic effects of Aspergillus niger and Trichoderma harzianum spores reached 0.47- and 0.55-log within 10 min, respectively. The inactivation efficiency and the synergistic effect would be affected by disinfectant concentration, pH, and temperature. The combined inactivation caused more violent oxidative stimulation and more severe damage to the fungal spores than the individual inactivation based on the flow cytometry analysis and the scanning electron microscopy observation. The synergistic effect during the combined inactivation process was attributed to the generation of hydroxyl radicals by the reaction between ozone and chlorine and the promotion of chlorine penetration by the destruction of cell wall by ozone. The combined inactivation efficiency in natural water samples was reduced by 26.4-43.8% compared with that in PBS. The results of this study provided an efficient and feasible disinfection method for the control of fungi in drinking water.

The aggregation of Aspergillus spores and the impact on their inactivation by chlorine-based disinfectants.

The formation of fungal biofilm goes through some different states, including monodisperse state, aggregated state, germinated state, hyphal and biofilm. The aggregation of spores is a primary step of fungal biofilm development in aquatic systems. Previous studies on the inactivation of fungi were mostly performed in the monodisperse state of fungal spores and biofilm state, however, the inactivation of aggregated fungal spores is still unclear. In this study, the aggregated characteristics of fungal spores (Aspergillus fumigatus and Aspergillus flavus) at different pH values were firstly studied, and the inactivation efficiency of fungal spores at different aggregation degree by chlorine-based disinfectants was also clarified. The results showed that the aggregation degree of Aspergillus fumigatus was the highest at pH 9.0 while it was the lowest at pH 5.0. Aggregation between fungal spores was mainly mediated by occasional adhesin-adhesin interactions and electrostatic interactions. Compared with monodisperse spores, fungal spores were more resistant to chlorine-based disinfectants with the increase of spore aggregation degree. The inactivation rate constants of Aspergillus fumigatus at 30% and 63% aggregation degree were 1.5- and 4-folds lower than that of monodisperse spores, respectively. The lower proportion of membrane damage and higher intracellular reactive oxygen species level for aggregated spores than monodisperse spores was observed according to the flow cytometric results after chlorine-based disinfectants treatment. The reasons for the lower inactivation efficiency of aggregated spores are as following: the protection of outer layer spores and signals between aggregates lead to the increase of resistance for aggregated spores. This study is meaningful for the control of the fungal spores at different states in water.

Evaluation of the vital viability and their application in fungal spores' disinfection with flow cytometry.

More attention was focused on fungi contamination in drinking water. Most researches about the inactivation of fungal spores has been conducted on disinfection efficiency and the leakage of intracellular substances. However, the specific structural damage of fungal spores treated by different disinfectants is poorly studied. In this study, the viability assessment methods of esterase activities and intracellular reactive oxygen species (ROS) were optimized, and the effects of chlorine-based disinfectants on fungal spores were evaluated by flow cytometry (FCM) and plating. The optimal staining conditions for esterase activity detection were as follows: fungal spores (106 cells/mL) were stained with 10 µM carboxyfluorescein diacetate and 50 mM ethylene diamine tetraacetic acid at 33 °C for 10 min (in dark). The optimal staining conditions for intracellular ROS detection were as follows: dihydroethidium (the final concentration of 2 µg/mL) was added into fungal suspensions (106 cells/mL), and then samples were incubated at 35 °C for 20 min (in dark). The cell culturability, membrane integrity, esterase activities, and intracellular ROS were examined to reveal the structural damage of fungal spores and underlying inactivation mechanisms. Disinfectants would cause the loss of the cell viability via five main steps: altered the morphology of fungal spores; increased the intracellular ROS levels; decreased the culturability, esterase activities and membrane integrity, thus leading to the irreversible death. It is appropriate to assess the effects of disinfectants on fungal spores and investigate their inactivation mechanisms using FCM.

Th1 and Th2 immune response in chronic hepatitis B patients during a long-term treatment with adefovir dipivoxil.

Adefovir dipivoxil treatment has significantly improved the outcome of chronic hepatitis B virus (HBV) infection. However, it remains largely unknown how immune system responds to the treatment. Chronic HBV patients were treated with adefovir dipivoxil and examined for serum HBV DNA loads, cytokines, and T helper (Th1) and 2 (Th2) cytokine producing T cells during 104 weeks of the treatment. Th1/Th2 cytokines producing T cells were significantly lower in chronic HBV patients as compared to normal individuals. Adefovir dipivoxil treatment led to the increase of Th1/Th2 cytokines producing T cells and serum cytokine levels in association with the decline of HVB DNA load. In contrast, Th1/Th2 cytokines producing T cells remained lower in one patient detected with adefovir dipivoxil resistant HBV A181T/V mutation. This study has established inverse correlation of the increase of Th1/Th2 immunity and the decline of HBV DNA load in chronic HBV patients during adefovir dipivoxil treatment.