No evidence of amplified Plasmodium falciparum plasmepsin II gene copy number in an area with artemisinin-resistant malaria along the China-Myanmar border.

BACKGROUND: The emergence and spread of artemisinin resistance in Plasmodium falciparum poses a threat to malaria eradication, including China's plan to eliminate malaria by 2020. Piperaquine (PPQ) resistance has emerged in Cambodia, compromising an important partner drug that is widely used in China in the form of dihydroartemisinin (DHA)-PPQ. Several mutations in a P. falciparum gene encoding a kelch protein on chromosome 13 (k13) are associated with artemisinin resistance and have arisen spread in the Great Mekong subregion, including the China-Myanmar border. Multiple copies of the plasmepsin II/III (pm2/3) genes, located on chromosome 14, have been shown to be associated with PPQ resistance. METHODS: The therapeutic efficacy of DHA-PPQ for the treatment of uncomplicated P. falciparum was evaluated along the China-Myanmar border from 2010 to 2014. The dry blood spots samples collected in the efficacy study prior DHA-PPQ treatment and from the local hospital by passive detection were used to amplify k13 and pm2. Polymorphisms within k13 were genotyped by capillary sequencing and pm2 copy number was quantified by relative-quantitative real-time polymerase chain reaction. Treatment outcome was evaluated with the World Health Organization protocol. A linear regression model was used to estimate the association between the day 3 positive rate and k13 mutation and the relationship of the pm2 copy number variants and k13 mutations. RESULTS: DHA-PPQ was effective for uncomplicated P. falciparum infection in Yunnan Province with cure rates > 95%. Twelve non synonymous mutations in the k13 domain were observed among the 268 samples with the prevalence of 44.0% and the predominant mutation was F446I with a prevalence of 32.8%. Only one sample was observed with multi-copies of pm2, including parasites with and without k13 mutations. The therapeutic efficacy of DHA-PPQ was > 95% along the China-Myanmar border, consistent with the lack of amplification of pm2. CONCLUSION: DHA-PPQ for uncomplicated P. falciparum infection still showed efficacy in an area with artemisinin-resistant malaria along the China-Myanmar border. There was no evidence to show PPQ resistance by clinical study and molecular markers survey. Continued monitoring of the parasite population using molecular markers will be important to track emergence and spread of resistance in this region.

In vivo monitoring of dihydroartemisinin-piperaquine sensitivity in Plasmodium falciparum along the China-Myanmar border of Yunnan Province, China from 2007 to 2013.

BACKGROUND: Artemisinin-based combination therapy (ACT) is the recommended first-line treatment of falciparum malaria in all endemic countries. Artemisinin resistance in Plasmodium falciparum has been confirmed in the Greater Mekong subregion (GMS). Dihydroartemisinin-piperaquine (DAPQ) is the most commonly used ACT in China. To understand the DAPQ sensitivity of P. falciparum, DAPQ resistance was monitored in vivo along the China-Myanmar border from 2007 to 2013. METHODS: Eligible patients with mono-infections of P. falciparum were recruited to this study after obtaining full informed consent. DAPQ tablets for different categories of kg body weight ranges were given once a day for three days. Patients were followed up for 42 days. Polymerase chain reaction (PCR) was conducted to distinguish between re-infection and recrudescence, to confirm the Plasmodium species. The data were entered and analysed by the Kaplan-Meier method. Treatment outcome was assessed according to the WHO recommended standards. RESULTS: 243 patients were completed valid follow-up. The fever clearance time (FCT) and asexual parasite clearance times (APCT) were, respectively, 36.5 ± 10.9 and 43.5 ± 11.8 hours, and there was an increasing trend of both FCT (F = 268.41, P < 0.0001) and APCT (F = 88.6, P < 0.0001) from 2007 to 2013. Eight (3.3%, 95% confidence interval, 1.4-6.4%) patients present parasitaemia on day three after medication; however they were spontaneous cure on day four. 241 (99.2%; 95% CI, 97.1-99.9%) of the patients were adequate clinical and parasitological response (ACPR) and the proportions of ACPR had not changed significantly from 2007 to 2013 (X(2) = 2.81, P = 0.7288). CONCLUSION: In terms of efficacy, DAPQ is still an effective treatment for falciparum malaria. DAPQ sensitivity in P. falciparum had not significantly changed along the China-Myanmar border of Yunnan Province, China. However more attentions should be given to becoming slower fever and parasite clearance.

[Establishment and Application of Multiplex PCR System for Detecting Four Human Plasmodium Species].

OBJECTIVE: To establish a multiplex PCR detection system for identifying 4 human Plasmodium species and evaluate its applicability. METHODS: The sequences of 18S rDNA gene of the 4 human Plasmodium species were compared using DNAman software, and 4 downstream primers were designed using Oligo 6.0 software, which targeted the region of variability between conserved regions 5 and 6 of the sequences. Using these primers, the specificity and sensitivity of the multiplex PCR system were evaluated, with plasmids containing the 18S rDNA gene sequence as a template. Further, a new nest PCR system (M-Nest) was established by combining the multiplex PCR system with the first-cycle genus-specific primer of the NP-1993 system. The sensitivities of the multiplex PCR system and the M-nest system were evaluated in serial dilutions of blood DNA samples from patients infected by P. falciparum and P. vivax. In addition, the NP-1993 and M-Nest systems were applied to screen the Plasmodium species in 307 blood samples from people returning to Guangxi from Ghana, a malaria epidemic area. And the NP-2002 and M-Nest systems were applied to re-check Plasmodium species in 66 blood samples collected in Guangxi from 2014 January to May, which were identified by microscopy to be infected mainly by P. ovale. RESULTS: The sizes of multiplex PCR products for P. falciparum, P. vivax, P. ovale, and P. malariae were 268 bp, 323 bp, 394 bp, and 446 bp, respectively, located in-between 50-bp DNA ladders. However, their melting curves had similar Tm values, thus could not be used to identify the 4 species. The minimum detection limits of P. falciparum, P. malariae, P. ovale, and P. vivax 18S rDNA gene by the multiplex PCR system were 5.58x102, 1.56x103, 1.66x103, and 1.80 x 10(2) copies/[A. The minimum detection limit of blood DNA from falciparum malaria patients by the multiplex PCR system was 1.43 x 10(2)-8.84 x 10(3) copies/p.1 or 5.10 x 10-4.92 x 10(2) parasites/µl, higher than that of P. vivax (17.4-69.1 copies/L or 13.5-83.2 parasites/p). Compared with this multiples PCR system, The M-Nest system further reduced the minimum detection limit of Plasmodium by 10-100 folds. Further, the M-Nest and NP-1993 systems reached inconsistent detection results in 307 blood samples from people returned to from Ghana; the former detected 2 cases of P. ovale infection while the latter failed. In addition, the NP-2002 and M-Nest systems came to the same results in re-checking Plasmodium species in the 66 blood samples. CONCLUSION: The established multiplex PCR system can identify 4 human Plasmodium species simultaneously and has good applicability in practice.

[Opportunities, Challenges and Strategies for Malaria Elimination in the Tibet Autonomous Region of China].

This review summarizes the endemic characteristics and influencing factors of malaria in the Tibet Autonomous Region of China, as well as the opportunities and challenges for malaria elimination in this region. Recommendations on strategies for malaria control are also made.

Monitoring Plasmodium vivax chloroquine sensitivity along China-Myanmar border of Yunnan Province, China during 2008-2013.

BACKGROUND: Plasmodium vivax is the most widespread of the malaria parasites infecting human hosts. In malaria-eliminating settings, both imported and local malaria predominantly occurs in border areas, and most of them are P. vivax. Chloroquine (CQ) is the first-line drug for P. vivax treatment in China. To understand CQ sensitivity in P. vivax, in vivo monitoring of CQ resistance was conducted along the China-Myanmar border from 2008 to 2013. METHODS: Eligible patients with mono-infections of P. vivax were recruited to this study after obtaining full informed consent. CQ tablets for different categories of kg body weight ranges were given once a day for three days. Patients were followed up for 28 days. PCR was conducted to distinguish between re-infection and recrudescence, to confirm the Plasmodium species. The data were entered and analysed by the Kaplan-Meier method. Treatment outcome and sensitivity were classified according to the WHO recommended standards. RESULTS: 603 patients were completed valid follow-up. The fever clearance time and asexual parasite clearance times were, respectively, 22.2 ± 10.2 and 38.1 ± 12.6 hours. 594 (98.5%) patients were adequate clinical and parasitological response (ACPR), and nine (1.5%) patients, who were late clinical failure (LCF) or resistant response level I (RI), were imported from the neighbouring districts of Myanmar. CONCLUSION: In terms of efficacy, CQ is still effective for vivax malaria treatment. Plasmodium vivax CQ sensitivity had not significantly changed along the China-Myanmar border of Yunnan Province, China.

Application of Auto-regressive Linear Model in Understanding the Effect of Climate on Malaria Vectors Dynamics in the Three Gorges Reservoir.

It is important to understand the dynamics of malaria vectors in implementing malaria control strategies. Six villages were selected from different sections in the Three Gorges Reservoir for exploring the relationship between the climatic factors and its malaria vector density from 1997 to 2007 using the auto-regressive linear model regression method. The result indicated that both temperature and precipitation were better modeled as quadratic rather than linearly related to the density of Anopheles sinensis.

[Overviews on Plasmodium ovale wallikeri and its genetic detection system].

In recent years, some microscopy-positive but nest-PCR negative (based on small subunit ribosomal RNA gene) ovale malaria cases were reported. The detection failure was due to the genetic variations between Plasomdoium ovale wallikeri and P. ovale curtisi. This paper reviews the discovery and validation processes of P. ovale wallikeri and the detection systems for identification of P. ovale.

Malaria transmission potential in the Three Gorges Reservoir of the Yangtze River, China.

OBJECTIVE: To define and evaluate the malaria transmission potential in the Yangtze River, following construction of the Three Gorges Reservoir. METHODS: Six villages, namely, Kaixian, Fengjie, Wanzhou, Fuling, Yubei, and Zigui were selected for investigating the malaria transmission potential in the reservoir. Transmission potential was estimated by mathematical modeling and evaluation of the local malaria situation. Factors that influenced the transmission potential were analyzed using Poisson regression analysis in combination with Grey Systematic Theory (based on evaluation by the Delphi method). RESULTS: Indirect fluorescent antibody data and the incidence of malaria in the local population were consistent with the malaria transmission potential calculated for the area. Multivariate Poisson regression analysis showed a statistically significant association between the riparian zone and the man-biting rate. CONCLUSION: The risk of a malaria epidemic can be forecasted using the malaria transmission potential parameters investigated here which was closely correlated with the riparian zone.

[Preliminary application of moving percentile method on surveillance and early-warning on visceral leishmaniasis in endemic areas].

OBJECTIVE: To apply moving percentile method on surveillance and early-warning on visceral leishmaniasis in Kashgar Region and evaluate its effect. METHODS: Incidence data of visceral leishmaniasis in Kashgar Region were collected from the National Web-based Infectious Diseases Report System. Monthly detection was carried out by using moving percentile method. The 50th percentile (P50), 70th percentile (P70) and 90th percentile (P90) of historical baseline data were calculated for drawing a control chart, and P70 was adopted as the warning threshold to determine whether an epidemic would appear. If the number of cases in one month is higher than the corresponding P70 of historical baseline data, the warning signal will be generated. The sensitivity, specificity and positive predictive value were calculated for the evaluation of early-warning effect. RESULTS: During the study period, 61.0% cases were reported in the year of 2008 and 2009, the incidence peak was from September to December, accounting for 51.9%, and infants under 3 years old were the population most threatened by visceral leishmaniasis, accounting for 62.7%. A total of 58 detections were performed, and 17 warning signals were generated by the threshold on P70. Among them, the numbers of cases in 9 detections were higher than the corresponding P90 of historical-baseline data. Based on the actually epidemic status of visceral leishmaniasis in study period, according to the threshold on P70, a total of 10 warning signals of 11 epidemics were detected, and the sensitivity of the warning model was 90.9% (10/11). 7 wrongly signals of 47 non-epidemics were detected, and the specificity was 85.1% (40/47). 10 of 17 signals were proved to be correct, and the positive predictive value was 58.8% (10/17). CONCLUSION: The moving percentile method can effectively perform surveillance and early-warning on visceral leishmaniasis in Kashgar Region.

Chemotherapy-based control of ascariasis and hookworm in highly endemic areas of China: field observations and a modeling analysis.

OBJECTIVE: Our objective was to systematically evaluate chemotherapy-based control of ascariasis and hookworm infection and make predictions of the effectiveness of repeated mass treatment at different levels of coverage in highly endemic areas of China. METHODS: Field surveys were carried out to acquire the ascariasis and hookworm prevalence and intensity (mean worm burden) at baseline, one month and one year later. We calculated model parameters based on the survey data, then incorporated them into a quantitative framework to predict the prevalence and intensity one year later. Sensitivity analyses were performed to assess the influence of the chemotherapy measures on prevalence and intensity, and model simulations were performed to evaluate the feasibility of achieving the proposed transmission control criteria under different chemotherapy measures. RESULTS: The predicted prevalence and intensity one year from baseline were within the 95% confidence interval of actual values. As treatment frequency or coverage increased, the prevalence and intensity decreased. Model simulations show that many rounds of treatment are needed to maintain the prevalence at a low level in highly endemic areas of China. CONCLUSION: We should select different combinations of treatment frequency, coverage and drug efficacy according to available resources and practical attainable conditions. Mathematical modeling could be used to help optimize the chemotherapeutic scheme aiming at specific parasitic species and areas, and to direct the establishment of soil-transmitted helminthiasis control criteria in China.

[Study on the correlation between land use and cover change and malaria transmission in the areas along the Yellow River and Huai River].

OBJECTIVES: To study the major ecologic factors influencing the re-emergence of malaria in the areas along the Yellow River and Huai River by analyzing the relationship between the malaria incidence and the land use and cover change (LUCC). METHODS: The data of annual parasite incidence (API) and LUCC in the counties of Huaiyuan, Yongcheng and Tong ai in 1990-2006 were collected retrospectively. Considering the hysteresis effect of LUCC played on malaria transmission, analysis of LUCC in these counties were conducted based on the data of 1990-1995 and 1996-2000, while the API data in 1996-2000 and 2001-2005 were used to analyze the trends and changes of malaria incidence. The correlation and trend analyses were conducted between LUCC and malaria incidence change in the periods. RESULTS: The LUCC in 1990-1995 in Tongbai County was significant, being 3 265.79% in the farmland and -90.42% in paddy field. The increase of malaria incidence in Tongbai was also significant in 1996-2000, the change rate was 2 799.70%, showing a positive correlation. The LUCC of other 2 counties in 1990-1995 was -0.27% and -0.78%, respectively, while the rate of malaria incidence change in 1996-2000 was 206.25% and 0.00%. The LUCC of the 2 counties in 1996-2000 was -0.08% and -0.50%, while the rate of malaria incidence change in 2001-2005 was 153.22% and 2 500.00%, respectively. It indicated that there was no significant relationship between malaria re-emergence and LUCC in Huaiyuan and Yongcheng Counties. CONCLUSION: It seems that the LUCC shows no significant impact on malaria re-emergence in areas along the Yellow River and Huai River since 2001, though there was a positive correlation between the two in Tonghai County in the late 1990s.

Establishing and applying an adaptive strategy and approach to eliminating malaria: practice and lessons learnt from China from 2011 to 2020.

ABSTRACTOn 30 June 2021, China was certified malaria-free by the World Health Organization. In this study, the evolution, performance, outcomes, and impact of China's adaptive strategy and approach for malaria elimination from 2011 to 2020 were analysed using 10-year data. The strategy and approach focused on timely detection and rapid responses to individual cases and foci. Indigenous cases declined from 1,308 in 2011 to 36 in 2015, and the last one was reported from Yunnan Province in April 2016, although thousands of imported cases still occur annually. The "1-3-7" approach was implemented successfully between 2013 and 2020, with 100% of cases reported within 24 h, 94.5% of cases investigated within three days of case reporting, and 93.4% of foci responses performed within seven days. Additionally, 81.6% of patients attended the first healthcare visit within 1-3 days of onset and 58.4% were diagnosed as malaria within three days of onset, in 2017-2020. The adaptive strategy and approach, along with their universal implementation, are most critical in malaria elimination. In addition to strengthening surveillance on drug resistance and vectors and border malaria collaboration, a further adapted three-step strategy and the corresponding "3-3-7" model are recommended to address the risks of re-transmission and death by imported cases after elimination. China's successful practice and lessons learnt through long-term efforts provide a reference for countries moving towards elimination.

[Investigation on Anopheles species and their composition in villages at different altitudes of Motuo County, Tibet Autonomous Region].

OBJECTIVE: To study the anopheline species and composition in villages at different altitudes, Muotuo County. METHODS: Six villages with different altitudes were selected as the investigation spots, i.e. Gande, Zhucun, Damu, Motuo, Didong and Beibeng with an altitude 1 966 m, 1 510 m, 1 408 m, 1 178 m, 853 m and 831 m, respectively. Human-baited net traps, cow-baited traps and light traps were set up to collect adult mosquitoes. The trapped mosquitoes were counted and identified according to morphological criteria. Following the classification, the mosquitoes were killed by chloroform and dried on silica-gel, and transported to the laboratory where they were stored at -20 degrees C. Species of Anopheles maculatus complex were identified with multiple PCR method. RESULTS: 5 410 anopheline mosquitoes were collected. Two mosquitoes were captured in high altitude village, one was Anopheles gigas bailieyi, while the other was damaged and unable to identify. There were 541 (36.9%) An. pseudowillmori, 906 An. willmori (61.7%) and 21 An. peditaeniatus (1.4%) collected in semi-high altitude villages; 260 (76.3%) An. pseudowillmori, 2 An. willmori (0.6%) and 79 An. peditaeniatus (23.2%) trapped in middle altitude village; and 3265 (90.7%) An. pseudowillmori, 19 An. willmore (0.5%) and 315 An. peditaeniatus (8.8%) trapped in low altitude villages. CONCLUSION: An. pseudowillmori, An. willmore and An. peditaeniatus make the main anopheline composition. The proportion of An. willmore is higher than An. pseudowillmori in semi-high altitude villages, while An. pseudowillmori take the absolute predominance in middle and low altitude villages.

Quantitative Microbial Risk Assessment of Cryptosporidium and Giardia in Public Drinking Water in China.

We aimed to assess the risks of Cryptosporidium and Giardia infections associated with drinking water for local residents, based on a quantitative microbial risk assessment, in three densely populated regions of China. In total, 45 source water samples and 45 treated water samples were collected from June to December 2014. Five Cryptosporidium-positive samples and 5 Giardia-positive samples were found. The annual probability of infection for individuals in Jintan (6.27 × 10 -4-2.05 × 10 -3 for Cryptosporidium and 7.18 × 10 -4-2.32 × 10 -3 for Giardia), Ezhou (6.27 × 10 -4-1.10 × 10 -2 for Cryptosporidium and 3.65 × 10 -4-1.20 × 10 -3 for Giardia), and Binyang (3.79 × 10 -4-1.25 × 10 -3 for Cryptosporidium) exceeded the tolerable risk of infection of 10 -4 set by the United States Environmental Protection Agency. Moreover, the corresponding disease burdens of cryptosporidiosis and giardiasis, due to direct drinking and residual water in these regions, exceeded the threshold of 10 -6 disability-adjusted life years per person per year set by the World Health Organization. These results provide insights into strategies to improve the safety of drinking water.

[Research progress on midgut bacteria of anopheline mosquitoes and their application in malaria control].

Midgut bacteria in anopheline mosquitoes could control the mosquito populations and block Plasmodium development. The article summarizes the role of bacteria in the mosquito nutrition, the midgut bacterial flora in anopheline mosquitoes and their application in disease control, transstadial transfer of bacteria within mosquitoes, the molecular immune responses of the mosquito to bacteria and the feasibility of applying bacteria to control malaria.

[The midgut bacterial flora in lab-reared Anopheles sinensis].

OBJECTIVE: To screen gram-negative bacteria in the midguts of lab-reared Anopheles sinensis at 3 development stages (larva, unfed adult and engorged adult). METHODS: Twenty engorged adult An. sinensis mosquitoes were dissected, and bacteria were isolated from their midguts by plate cultivation. The bacteria were identified by using 16S rDNA V3 sequence and dyed with Gram-reaction solutions. Using denaturing gradient gel electrophoresis (DGGE), 16S rDNA clone libraries from common bacteria in the midguts of lab-reared An. sinensis at 3 development stages (20 engorged adults, 10 larva and 50 unfed adults) were constructed. Target Gram-negative bacteria were screened by phylogenetic analysis of all 16S rDNA V3 sequences. RESULTS: In engorged adult female An. sinensis, 16S rRNA sequencing placed the 28 screened bacterial colonies with their closest matches into 5 major groups. Except Gram-positive Brevibacillus sp., the other four, Aeromonas sp., Comamonas sp., Elizabethkingia sp., and A4 were Gram-negative. DGGE analysis showed that 4 distinct bands existed in 3 development stages of An. sinensis. 16S rDNA V3 segments revealed 5 species of bacteria which were Aeromonas sp., A4, Pseudomonas sp., Leucobacter sp., and one unclassified bacterium. Compared with 16S rDNA V3 region of Aeromonas sp. (GQ301543) and A4(FJ8701127), the matching 16S rDNA segments of midgut bacteria of engorged adults showed a nucleotide sequence similarity of 100%. CONCLUSION: Aeromonas sp. and A4 were the common midgut Gram-negative bacteria of An. sinensis at 3 development stages.

[Intelligence level and characteristics of cognitive structure in school-age children infected with soil-transmitted helminthes].

OBJECTIVE: To investigate the intelligence level and characteristics of cognitive structure of school-age children infected with soil-transmitted helminthes (STH) (hookworm, Ascaris lumbricoides and Trichuris trichiura). METHODS: 1 031 children of 9 to 12 years old from Baishui village of Guangxi, Longzhou and Xinzhu villages of Hainan were selected for the survey. Stool samples from children were collected and examined by modified Kato-Katz method. 77 children infected with only one species (hookworm, A. lumbricoides, or T. trichiura) were used as case group and 39 healthy children as control. The cognitive ability for children were assessed by WISC-LV. RESULTS: A total of 378 cases with STH infection was found from 1 031 children, and the overall prevalence of STH infection was 36.7%. The prevalence of moderate and heavy STH infections was 16.8% (173/1 031). The verbal comprehension (VCI), working memory (WMI), processing speed (PSI) and full scale IQ (FSIQ) indices of case group were lower than those of control group (P < 0.05). The VCI (97.2), WMI (84.6), PSI (91.1) and FSIQ (95.4) indices in moderate/heavy infection group were lower than those in light infection group (99.2, 103.3, 88.3 and 99.1, respectively) and control group (102.0, 104.2, 91.9 and 100.1, respectively) (P < 0.05). The WMI index in hookworm infection group (83.7) was lower than that in A. lumbricoides (87.6) or T. trichiura (88.3) infection groups (P < 0.05). The PSI index of T. trichiura infection group (92.8) was lower than that of A. lumbricoides (97.3) or hookworm (94.4) infection groups (P < 0.05). CONCLUSION: The total intelligence level of STH infection children is in a normal range, or on borderline. The low VCI, WMI, PSI indices have been found in moderate/heavy STH infection children.

[Dendritic cell function in response to schistosome infection].

Dendritic cells (DC) are uniquely specialised for both antigen acquisition and presentation, linking innate and adaptive immunity. Great progress has been made in DC biology and function recently. In the field of schistosomiasis, studies have revealed that a DC phenotype is quite distinct to that of conventional activation. Understanding the interaction between schistosomes and DC is therefore not only addressing fundamental questions of DC biology and immunity to multicellular parasites but also opening the way to therapeutic manipulation of the immune system.

[Analysis on the award-winning research achievements by the National Institute of Parasitic Diseases].

OBJECTIVE: To evaluate the predominance, characteristic and shortage of the scientific activities at the National Institute of Parasitic Diseases (NIPD), through analysis on its award-winning research achievements. METHODS: Information was collected on the award-winning research achievements by the Institute in the last 45 years. Time trend, disease category, subject, award type, award grade, award level and project property of the award-winning research achievements were analyzed by using SPSS11.0 software. RESULTS: Totally, 78 research achievements were awarded 128 times in the last 45 years. 43.6% awards were at province and ministry level; 23.1% awards were at nation level. Awards involved in schistosomiasis, malaria, and kala-azar accounted for 33.3%, 28.2% and 12.8%, respectively. As for subject, preventive medicine, pharmacy and biology accounted for 28.2%, 18.0% and 16.7%, respectively; 82.1% of the awards belonged to applied researches. CONCLUSION: NIPD has a strong capacity in scientific research. To get more achievements in future, it is essential to closely integrate the experiment research to disease control.

[Protective efficacy induced by dendritic cells pulsed with GST in combination with CpG oligodeoxynucleotide against Schistosoma japonicum infection].

OBJECTIVE: To study the protective effects on the infection of Schistosoma japonicum in C57BL/6 mice induced by dendritic cells DCs pulsed with GST in combination with CpG oligodeoxynucleotide. METHODS: GST was purified and used to stimulate DC2.4 cell line. Antigen loading was analyzed by immunofluorescence method. Thirty-five C57BL/6 mice were divided into seven groups(5 mice per group). Mice in groups A, B, C, D and E were immunized subcutaneously with DCs, DCs treated with PSA, DCs pulsed with GST, DCs stimulated with GST+CpG ODN, DCs stimulated with CpG ODN, respectively. For the above five groups, each mouse received 100 microl cell suspension at the density of 10(7)/ml subcutaneously for three times at 2-week intervals. Each mouse of group F was immunized subcutaneously with 50 microg GST formulated in complete Freund's adjuvant first, and 50 microg, 10 microg GST respectively in incomplete Freund's adjuvant for the last two doses. Group G received PBS and served as control. Serum samples were collected 10 days after the final immunization, and were analyzed for specific antibodies by ELISA. At two weeks after the final immunization, each mouse were challenged by 30 +/- 1 cercariae of S. japonicum. Six weeks after infection the mice were sacrificed, and number of worms was counted. RESULTS: Light green fluorescence was observed in dendritic cells under the fluoroscope after pulsing with GST which indicated the protein loaded dendritic cells. The IgG level in groups C, D and F was 0.555 2 +/- 0.078 9, 0.715 0 +/- 0.052 3, and 2.127 0 +/- 0.411 5, respectively, all higher than that of group G (P < 0.05). The worm reduction rate of group D was 53.3%, followed by group F (24.0%) and group C (21.3%). There was no significantly difference in the worm reduction rate between group D and groups F and C (P > 0.05). CONCLUSION: Dendritic cells pulsed with GST in combination with CpG oligodeoxynucleotide induce significant immunoprotection against the infection of Schistosoma japonicum.