RESUMEN
The neurotropic parasite Toxoplasma gondii (T. gondii) infection can change the behavior of rodents and cause neuropsychological symptoms in humans, which may be related to the change in neurotransmitter dopamine in the host brain caused by T. gondii infection. T. gondii tyrosine hydroxylase (TgTH) is an important factor in increasing the neurotransmitter dopamine in the host brain. In this study, the enzyme activity of TgTH catalytic substrate for dopamine production and the molecular characteristics of TgTH were identified. In order to amplify the open reading frame (ORF), the designing of the specific primers for polymerase chain reaction (PCR) was on the basis of the TgTH sequence (GenBank Accession No. EU481510.1), which was inserted into pET-32a (+) for the expression of recombined TgTH (rTgTH). The sequence analysis indicated that the gene of TgTH directed the encoding of a 62.4-kDa protein consisting of 565 amino acid residues, which was predicted to have a high antigen index. The enzyme activity test showed that rTgTH and the soluble proteins extracted separately from T. gondii RH strain and PRU strain could catalyze the substrate to produce dopamine in a dose-dependent manner, and the optimum catalytic temperature was 37 °C. The result of the Western Blotting assay revealed that the rTgTH and the native TgTH extracted from somatic of T. gondii RH tachyzoite were successfully detected by the sera of mice infected with T. gondii and the rat serum after rTgTH immune, respectively. Immunofluorescence analysis using antibody against rTgTH demonstrated that the protein was expressed and located on the surface of T. gondii RH tachyzoite. Freund's adjuvant was used to emulsify the rTgTH, which was subsequently applied to BALB/c mouse immune thrice on week 0, week 2, and week 4, respectively. The result of the animal challenge experiments showed an integral increase in IgG, IgG2a, IgG1, and IFN-γ, IL-4, and IL17 were as well significantly increased, and that the rTgTH vaccinated animals apparently had a prolonged survival time (14.30 ± 2.41) after infection with the RH strain of T. gondii compared with that of the non-vaccinated control animals, which died within 11 days. Additionally, in the rTgTH vaccination group, the number of brain cysts (1275 ± 224) significantly decreased (p < 0.05) compared to the blank control group (2375 ± 883), and the size of the brain cysts in the animals immunized with rTgTH vaccination was remarkably smaller than that of the control mice. All the findings prove that TgTH played an important role in increasing the neurotransmitter dopamine in the host brain and could be used as a vaccine candidate antigen to mediate cell-mediated and humoral immunity.
RESUMEN
The management and remediation of abandoned hydrocarbon-contaminated sites require detailed information on the distribution of contaminant plumes. In areas where groundwater is active, the formation of contaminant plumes is associated with hydrodynamics, the nature of the sedimentary layers, and the nature of the pollutants and the degradation process. A comprehensive survey is needed to determine this information. An abandoned hydrocarbon disposal site is located in an area where groundwater is very active. In the investigation of contaminant plumes, we combined the geophysical method with accurate geochemical analysis of subsoil and groundwater samples. Ground-penetrating radar and electrical resistivity tomography images of the electrical anomalies potentially originating from hydrocarbon pollution were used to select sites for subsurface sampling. Total petroleum hydrocarbons, total dissolved solids, and groundwater pH were measured. The results showed that the source zone had undergone long-term natural attenuation, and it was unable to continuously output organic matter to support the expansion of contaminant plumes. Low-resistivity anomalies and enhanced attenuation in the study area were caused by hydrocarbon degradation products and enhanced mineral weathering. Delineating the distribution of contaminant plumes in areas where the resistivity was below 15 Ω m. The distribution of the plume in the vertical direction was related to the hydrocarbon release history (release rate and volume) and was affected by fluctuations in the groundwater level. The contaminant plume moved very slowly along the direction of the hydraulic gradient and was in a basically stable state. The results showed that the combined application of the geoelectrical method and the geochemical method can effectively describe the distribution of underground contaminant plumes in an aged pollution site.