Comparative Study on Different Remediation Strategies Applied in Petroleum-Contaminated Soils.

Due to the increasing pollution by petroleum hydrocarbons (PHs), it is an important task to develop eco-friendly and highly efficient methods for remediating petroleum-contaminated soils. In this study, bioremediation technology was applied to remediate PHs contaminated soils, and the bacterial community structure and physicochemical characteristics of the soil treated using different bioremediation regimens were analyzed. Compared with the control condition (S0), the PHs removal efficiency of biostimulation (S2) and bioaugmentation (S3) was increased significantly. Combined biostimulation with bioaugmentation (S4) had the highest PHs removal efficiency, up to 60.14 ± 4.12%. Among all the selected remediation strategies (S1-S4, S1: soil moisture content: 25-30%), the bacterial alpha-diversity was higher than in S0. The genera Acinetobacter, Escherichia-Shigella, Bacteroides, Microbacterium, and Parabacteroides were found to greatly contribute to PHs' degradation. In the group S4, the PH-degraders and soil enzyme activity were higher than in the other remediation regimens, and these indices gradually decreased in the mid-to-later periods of all remediation tests. Additionally, the abundance of alkB and nah genes was increased by improving the environmental condition of the microorganism communities. Redundancy analysis (RDA) revealed that the total nitrogen (TN) and total phosphorus (TP) had a positive correlation with total PHs degradation. This study offers insights into the microbial community response to environmental factors during bioremediation, which shows a promoting effect in enhancing the efficiency of PHs remediation.

Effects of Funneliformis mosseae on Root Metabolites and Rhizosphere Soil Properties to Continuously-Cropped Soybean in the Potted-Experiments.

Continuous cropping in soybean is increasingly practiced in Heilongjiang Province, leading to substantial yield reductions and quality degradation. Arbuscular mycorrhizal fungi (AMF) are soil microorganisms that form mutualistic interactions with plant roots and can restore the plant rhizosphere microenvironment. In this study, two soybean lines (HN48 and HN66) were chosen as experimental materials, which were planted in different years of continuous cropping soybean soils and were inoculated or not with Funneliformis mosseae in potted-experiments. Ultimately, analysis of root tissue metabolome and root exudates, soil physicochemical properties, plant biomass, as well as rhizosphere soil properties in different experimental treatments, inoculated or not with F. mosseae, was performed. Experimental results showed that: (a) The disease index of soybean root rot was significantly lower in the treatment group than in the control group, and there were differences in disease index and the resistance effect of F. mosseae between the two cultivars; (b) compared with the control, the root tissue metabolome and root exudates remained unchanged, but there were changes in the relative amounts in the treatment group, and the abundant metabolites differed by soybean cultivar; (c) soybean biomass was significantly higher in the treatment group than in the control group, and the effect of F. mosseae on biomass differed with respect to the soybean cultivar; and (d) there were differences in the physiochemical indexes of soybean rhizosphere soil between the treatment and control groups, and the repairing effect of F. mosseae differed between the two cultivars. Therefore, F. mosseae can increase the biomass of continuously cropped soybean, improve the physicochemical properties of the rhizosphere soil, regulate the root metabolite profiles, and alleviate barriers to continuous cropping in potted-experiments of soybean.

[The effect of aluminum adjuvant and immunization schedule on immunogenicity of Sabin inactivated poliovirus vaccine].

OBJECTIVE: To study the effect of aluminume adjuvant and immunization schedule on immunogenicity of Sabin inactivated poliovirus vaccine. METHODS: Four batches of Sabin IPV were produced by different concentrations of type 1, 2, and 3 poliovirus and administrated on three-dose schedule at 0, 1, 2 months and 0, 2, 4 months on rats. Serum samples were collected one month after each dose and neutralizing antibody titers against three types poliovirus were determined by micro-neutralization assay. RESULTS: The GMTs of neutralizing antibodies against three types poliovirus increased significantly and the seropositivity rates were 100% in all groups after 3 doses. There was no significant difference between two immunization schedules, and the 0, 2, 4 month schedule could induce higher level neutralizing antibody compared to the 0, 1, 2 month schedule. The groups with aluminum adjuvant could induce higher level neutralizing antibody compared to the groups without adjuvant. CONCLUSION: Aluminum djuvant and immunization schedule could improve the immunogenicity of Sabin IPV.

[Hydroacoustic assessment of fish resources in reservoirs with different fishery management].

By using Biosonics DT-X echosounder (208 kHz), a hydroacoustic investigation was conducted on the fish resources in three mesotrophic reservoirs (Daxi Reservoir, Shahe Reservoir, and Jinniushan Reservoir) with different fishery management in late autumn and early winter 2011, and a GIS model was constructed to assess the fish resource distribution in the reservoirs. No significant difference was observed in the average size of fish in the three reservoirs, but the distribution curves of fish target strength (TS) showed that the fish size distribution proportion was different, and had close correlation with fishery management. The fish density in Daxi Reservoir (averagely 0.0183 ind x m(-3)) had no significant difference with that in Shahe Reservoir (averagely 0.0124 ind x m(-3)), but the fish density in Jinniushan Reservoir (averagely 0.0085 ind x m(-3)) was significantly lower than that in Daxi and Shahe reservoirs. There was no significant correlation between the horizontal distribution of fish density and the water depth in the three reservoirs. The fish in the three reservoirs were in group distribution, and fish aggregation was found in Daxi and Shahe reservoirs. In the three reservoirs, the fish biomass was the highest in downstream, and there was a greater biomass near the dam, except in Shahe Reservoir which was affected by fish catch activity. Based on the raster data obtained from GIS fish distribution model and the surface water area of each grid, the total amount of fish resources with the TS > -60 dB in the Daxi, Shahe and Jinniushan reservoirs was estimated as about 480000, 610000 and 520000 individuals, and that with the TS > -40 dB was about 50400, 52900 and 90700 individuals, respectively.

[Experimental study on the environmental fate of nitrogen in snail-macrophyte ecosystem for water purification].

A snail-macrophyte simulation system was built and isotope tracer technique was adopted to study the environmental fate of nitrogen in snail-macrophyte purification system, the results showed that: Vallisneria spiralis increased its wet weight by 580% and its number by 6.6 ramets, moreover, Vallisneria spiralis absorbed 1.07% 15N by the roots and 7.74% by stems and leaves, while Bellamya only absorbed 0.06%. And 5.73% 15N was retained in the sediment. Through analyzing of the results, it indicated that: in such simulation system, sediment was the main nutrition source for the growth of Vallisneria spiralis, which absorbed only few dissolved nitrogen from water; ammonium nitrogen in water was transformed mainly in the sediment-water interface, most of which was absorbed by Vallisneria spiralis, a small amount was removed through nitrification and denitrification, and the rest was kept by sediment; Vallisneria spiralis was final vector for removing nitrogen in the system, and Bellamya could also boost the growth of Vallisneria spiralis and strengthen the processes of nitrification and denitrification, thus promoting the nitrogen removal from the system indirectly. So, during the period of culture, rational allocation of snail-macrophyte structure in different stages plays an important role in controlling water quality in ponds.

[Immunogenicity of sabin inactivated poliovirus vaccine induced by diphtheria-tetanus-acellular pertussis and Sabin inactivated poliovirus combined vaccine].

OBJECTIVE: In order to search the preparation process and optimazing dosage ratio of adsorbed diphtheria-tetanus-acellular pertussis and sabin inactivated poliovirus combined vaccine (DTaP-sIPV), the neutralizing antibody titers of IPV induced by different concentration of DTaP-sIPV were investigated on rats. METHODS: Two batches of DTaP-sLPV were produced using different concentration of sIPV and the quality control was carried. Together with sabin-IPV and DTaP-wIPV ( boostrix-polio, GSK, Belgium) as control group, the DTaP-sIPV were administrated on three-dose schedule at 0, 1, 2 month on rats. Serum sample were collected 30 days after each dose and neutralizing antibody titers against three types poliovirus were determined using micro-neutralization test. RESULTS: Two batches of prepared DTaP-sIPV and control sLPV were according to the requirement of Chinese Pharmacopoeia (Volume III, 2005 edition) and showed good stability. The seropositivity rates were 100% for sabin inactivated poliovirus antigen in all groups. The GMTs (Geometric mean titers) of neutralizing antibodies against three types poliovirus increased. CONCLUSION: The prepared DTaP-sIPV was safe, stable and effective and could induced high level neutralizing antibody against poliovirus on rats.

[Preparation of vero cell-adapted influenza H5N1 virus strain by genetic reassortment].

OBJECTIVE: To prepare Vero cell-adapted influenza H5N1 virus strain by Genetic Reassortment and produce influenza H5N1 vaccine using Vero cell as a substrate. METHODS: Embryonated specific pathogen-free (SPF) hen's eggs and Vero cells were co-infected with Vero cell-adapted influenza virus A/Yunnan/1/2005 Va(H3N2) and A/Anhui/1/2005 (H5N1) via reverse genetics. The reassortant was screened with goat antibody against strain A/Yunnan/1/2005 Va(H3N2) and identified for subtype by hemagglutination-inhibition (HI) assays and gene analysis of HA and NA. RESULTS: A Vero cell-adapted influenza H5N1 virus strain was obtained, and there was no significant difference in serum antibody titers of monovalent inactivated vaccine reassorted before and after (F = 0.857, P > 0.05). CONCLUSION: The Vero cell-adapted influenza virus of epidemic strain may be reassortment between Vero cell-adapted and epidemic strains.

Screening of a high growth influenza B virus strain in Vero cells.

Due to the insufficient supply of embryonated chicken eggs, the preparation of large quantities of inactivated influenza vaccines will require an alternative virus culture system after the emergence or reemergence of a pandemic influenza virus. The Vero cell is one of the ideal options since it was used for producing many kinds of human vaccines. However, most of the influenza viruses can not grow well in Vero cells. To develop a new influenza vaccine with Vero cells as a substrate, the virus needs to adapt to this cell substrate to maintain high growth characteristics. By serial passages in Vero cells, the B/Yunnan/2/2005va (B) strain was successfully adapted to Vero cells, with the hemagglutination titer (HAT) of the virus reaching 1:512. The high growth characteristic of this strain is stable up to 21 passages. The strain was identified by hemagglutination inhibition (HAI) test and sequencing respectively; the HA1 gene sequence of the virus was cloned and analyzed. The screening and establishment of high growth B virus provides an important tool for influenza vaccine production in Vero cells.

[Effective inactivatian test of inactivated hepatitis A vaccine using integrated cell culture/strand-specific reverse transcriptase-polymerase chain reaction].

OBJECTIVE: To establish an quick, sensitive and specific assay for effective inactivatian test of inactivated hepatitis A vaccine. METHODS: effective inactivatian test of inactivated hepatitis A vaccine were carried out using integrated cell culture/strand-specific RT-PCR (ICC/strand-specific RT-PCR) assay compared with traditional ELISA and nest RT-PCR assay. RESULTS: all the samples were infectious negative detecting by both ICC/ strand-specific RT-PCR and ELISA assay,while some samples appeared false positive detecting by nest RT-PCR. CONCLUSION: ICC/strand-specific RT-PCR assay is a novel, rapid, sensitive and reliable method for effective inactivatian test of inactivated hepatitis A vaccine. Shorting detection period largely, this assay may be used as an alternative method for routine inactivated hepatitis A vaccines test.

[Research on an improved Lowry method to determine content of protein in Sabin IPV].

OBJECTIVE: To establish a method for the content determination of protein in Sabin IPV. METHODS: Using lowry method combined with being precipitated by trichloroacetic acid to determine the content of protein in Sabin IPV. Changing different conditions to optimize the experiment to establish a improved lowry method. And the sample recovery test was also conducted. RESULTS: The method can exclude the interference of free aminoacid, phenols and some other additives. The calibration curve was in good linearity of protein within the range of 2.5 microg/ml-40 Microg/ml, r = 0.9998. Under the best conditions, the mean recovery was 95.32%, the CV in a batch and between batches were both < 10%. CONCLUSION: The method can be used to determine the micro content of protein in vaccines.

[Effect of 2-phenoxyethanol on potency of Sabin inactivated poliomyelitis vaccine and its safety].

OBJECTIVE: To investigate the effect of 2-phenoxyethanol on potency of Sabin inactivated poliomyelitis vaccine (IPV). METHODS: Sabin IPV samples containing 5 mg or 7 mg 2-phenoxyethanol each dosage respectively were placed separately at 4 degrees C, 37 degrees C for 2 days and 7 days. D-antigen contents were tested with ELISA method. Then neutralizing antibodies in mice and guinea pigs were detected. The safety experiment was performed according to unusual toxicity test of China requirement for biological product. RESULTS: After addition of 2-phenoxyethanol, the I, II, and III D-antigen contents of Sabin IPV did not change. The antibody levels in mice and guinea pigs were not different between experimental group and control group. Animals were safe during observation period. CONCLUSION: 2-Phenoxyethanol had no effect on potency and safety of Sabin IPV. It can be used as antiseptic for Sabin IPV.

A novel process for production of hepatitis A virus in Vero cells grown on microcarriers in bioreactor.

AIM: To develop a novel process for production of HAV in Vero cells grown on microcarriers in a bioreactor. METHODS: Vero cells infected with HAV strain W were seeded at an initial density of 1 x 10(5) cells/mL into a 7-L bioreactor containing Cytodex-I microcarriers. During the stage of cell proliferation, the following conditions were applied: pH 7.2 +/- 0.2, temperature 37 +/- 0.2 degrees, dissolved oxygen 40% of air saturation and agitation rate 40 r/min. After the stage of virus culture started, the culture conditions were altered to pH 7.2 +/- 0.2, temperature 35 +/- 0.2 degrees, dissolved oxygen 25% of air saturation, agitation rate 50 r/min and perfusion of fresh medium at a flux of 20 mL/h. During the course of fermentation, cell density, HAV antigen titre, glucose, lactate and ammonia levels were monitored. A control experiment using conventional static culture was conducted in the T150 flask. RESULTS: After a 28-d cultivation, cell density increased to 14.0 x 10(6) cells/mL in the bioreactor, 5.6 x 10(9) viable cells and 4,000 mL virus suspension with a titre of 1:64 were harvested. The viral antigen output per cell unit in the bioreactor was 3-fold higher than that in the T150 flask. Meanwhile the metabolic mode of Vero cells did not change after the infection with HAV strain W. CONCLUSION: The process for production of HAV in Vero cells grown on microcarriers in a bioreactor is a novel, efficient and practical way to obtain virus antigen for vaccine purpose. This approach produces more cells and HAV antigen than the conventional static culture. With further improvement, it is possible to be used for the production of hepatitis A vaccine.

[Immunogenecity of combined hepatitis A and B vaccine].

OBJECTIVE: To observe the immunogenicity of combined hepatitis A and B vaccine (HAB). METHODS: The combined HAB vaccine was prepared and different concentrations of HAB were administered on mice in week 0, 4 and 24, and then we tested the antibodies to both hepatitis A virus and B virus. After the first injection, we tested the hepatitis A antigen-induced and hepatitis B surface antigen-induced stimulation indices in spleen monocyte as well as changes of CD4+ and CD8+ cell numbers. RESULTS: The serum antibody positive rates were 100% in all three groups, and the antibody induced by HAB vaccine were earlier than by monovalent vaccine. The hepatitis A antibody and hepatitis B surface antibody titers after the combined vaccine inoculation were not significantly higher than those after the monovalent vaccine inoculation. On the other hand, after the first injection of the combined vaccine, the hepatitis A antigen-induced and hepatitis B surface antigen-induced stimulation indices in spleen monocyte were detected. The numbers of CD4+ and CD8+ cells increased. CONCLUSIONS: HAB vaccine has reliable immunogenicity.