Seroprevalence and genetic characterization of Toxoplasma gondii in cancer patients in Anhui Province, Eastern China.

BACKGROUND: Recent studies have indicated the predominance of Toxoplasma gondii genotype Chinese 1 in animals in China. However, little is known of the genetic features of the parasite in humans. This study aims to determine the prevalence of anti-T. gondii antibodies based on which the genetic character of the parasite was identified in cancer patients in China. METHODS: A total of 1014 serum samples with malignant neoplasms were collected from six tertiary-care hospitals (HAUCM, APH, HAMU, XAH, FHH and HBMC) from January, 2012 to August, 2013. Antibodies against T. gondii were examined by enzyme-linked immunosorbent assay (ELISA). Blood samples were subsequently used for PCR assay to detect T. gondii DNA (gra6). The DNA positive samples were subjected to genotyping using a multiplex multilocus nested PCR-RFLP at 10 loci, including sag1, sag2, sag3, btub, gra6, l358, c22-8, c29-2, pk1 and apico. Samples from the patients were anonymous and only data with regard to age and gender was available at sample collection. RESULTS: Overall, 8.38% (85/1014) of the examined patients showed positive antibodies against T. gondii. Among them, 61 (6.02%) were seropositive only for IgG, 16 (1.58%) were only for IgM, and 8 (0.79%) were found to be positive for both IgG and IgM. The seroprevalence of antibodies to Toxoplasma ranged from 5.8% to 11.0%, without regional difference (χ(2) = 4.764, P = 0.445). No significant differences of the positive rates of T. gondii infection were noted in genders (male, 8.96%; female, 7.45%) (χ(2) = 0.707, P = 0.400) and in ages (χ(2) = 1.172, P = 0.947). Of 1014 DNA samples, 36 (3.55%) were positive for T. gondii by nested PCR at gra6 locus and nine gave rise to complete genotyping results. All samples with achieved PCR-RFLP genotyping showed a common genetic character of type Chinese 1 (ToxoDB#9). CONCLUSION: Seroprevalence of toxoplasmosis in immunosuppressed individuals is rarely reported in China and we presented a positive rate of 8.38% in cancer patients. Toxoplasma genomic DNA genotyping demonstrated a common genetic character of Chinese 1, indicating a possible pathogenic origin of animals in human infection.

[Kinetic distribution of cyst-forming Chinese isolate of Toxoplasma gondii in mice].

OBJECTIVE: To determine the kinetics of infection and cyst formation in CD1 mice following oral infection with cyst-forming Chinese isolate of Toxoplasma gondii TgCtwh1(genotype China 1, ToxoDB#9). METHODS: 50 CD1 female mice were obtained from specific pathogen-free (SPF) mouse colony in the Vital River Laboratories (VRL), Beijing. Mice were randomly divided into 10 groups each with 5 mice. All mice but control were peroral gavage infected with 50 cysts (1x10(4) bradyzoites) of TgCtwh1 isolate of T. gondii isolated from Wuhan, China. Cysts were isolated from the entire brain of mice infected with TgCtwh1 by density gradient centrifugation over Fycoll-paque plus. Animals were orally inoculated with cysts on day zero, and peripheral blood, lymph nodes, heart, liver, and brain of infected mice were collected on days 2, 4, 7, 10, 14, 21, 35, 50, and 72 post infection. Five mice were sacrificed by cervical dislocation under anesthesia at each time of collection, and the kinetic distribution was detected by fluorescence quantitative PCR and tissue inoculation into fresh mice. The cyst formation at various intervals after infection was also observed, as was the number of the cysts in brains and the cyst-forming rate. RESULTS: The body weight of the mice lessened (3.650 +/- 0.252)g post oral infection on day 7, and the weight was progressively decreased between day 10 [(1.730 +/- 0.017)g] and day 14 [(-0.390 +/- 0.554) g] after infection (P<0.05). In the brain tissue, cysts were first observed on day 21 post oral infection and the cyst-forming rate was 80%, and the average diameter of cysts was 20-40 microm. While on day 35 after infection, the cysts were formed in all infected mice(cyst-forming rate was 100%) and the average diameter was 50-60 microm. In chronic infection, DNA copies of parasites were first detected in blood, heart, liver and lymph node at 3.51 +/- 0.152, 4.100 +/- 0.198, 4.220 +/- 0.209 and 4.960 +/- 0.052 respectively on day 2, then in the brain on day 4 (3.800 +/- 0.154). During the early days of infection, the parasite burden in blood was progressively increased until days 7 (5.240 +/- 0.115) then gradually decreased and become undetectable on day 35. The burden of T. gondii in the heart and brain tissues increased significantly and reached their maximum on day 14 (5.640 +/- 0.214) and day 10 (5.790 +/- 0.060), respectively, and remained a stable level thereafter. Liver and lymph tissues reached their maximum on day 7 (5.310 +/- 0.038) and day 10 (6.200 +/- 0.152), then gradually decreased and become undetectable on day 50. CONCLUSION: The parasitemia in mice infected with T. gondii cyst-forming isolate lasts for 21 d at least, and cysts are detected in brain on day 21.