Table of Content

15 June 2024, Volume 36 Issue 3

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Progress of schistosomiasis control in People’s Republic of China in 2023

ZHANG Lijuan, HE Junyi, YANG Fan, DANG Hui, LI Yinlong, GUO Suying, LI Shizhen, CAO Chunli, XU Jing, LI Shizhu

2024, 36(3):  221-227. 

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To understand the progress of national schistosomiasis elimination program of China in 2023 and summarize the lessons and experiences, data on the endemic status of schistosomiasis and national schistosomiasis surveillance results in the People’s Republic of China were collected and analyzed at a national level. By the end of 2023, Shanghai Municipality, Zhejiang Province, Fujian Province, Guangdong Province and Guangxi Zhuang Autonomous Region continued to consolidate the achievements of schistosomiasis elimination, and Sichuan and Jiangsu provinces maintained the criteria of transmission interruption, while Yunnan and Hubei provinces were identified to achieve the criteria of transmission interruption in 2020, and Anhui, Jiangxi and Hunan provinces achieved the criteria of transmission interruption in 2023. A total of 451 counties (cites, districts) were found to be endemic for schistosomiasis in China in 2023, including 26 250 endemic villages covering 73 034 500 residents at risk of infections. Among the 451 endemic counties (cities, districts), 78.49% (354/451) achieved the criteria of schistosomiasis elimination and 21.51% (97/451) achieved the criteria of transmission interruption, respectively. In 2023, a total of 4 216 643 individuals received immunological tests, with 47 794 sero⁃positives identified, and a total of 184 216 individuals received parasitological examinations, with 4 egg⁃positives detected. A total of 27 768 cases with advanced schistosomiasis were documented in China by the end of 2023. In 2023, 539 548 bovines were raised in schistosomiasis⁃endemic areas of China, and 125 440 bovines received immunological tests, with 124 sero⁃positives detected, while no egg⁃positives were identified among the 133 508 bovines receiving parasitological examinations. In 2023, snail survey was performed at an area of 641 339.53 hm2 and 184 819.77 hm2 snail habitats were identified, including 51.53 hm2 emerging snail habitats and 642.25 hm2 reemerging snail habitats. In 2023, there were 20 198 schistosomiasis patients receiving praziquantel chemotherapy, and 598 183 person⁃time individuals and 283 954 herd⁃time bovines were given expanded chemotherapy. In 2023, snail control with chemical treatment was performed in 116 347.95 hm2 snail habitats, and the actual area of chemical treatment was 65 690.89 hm2, while environmental improvements were performed in snail habitats covering an area of 1 334.62 hm2. The national schistosomiasis surveillance results showed that the mean prevalence of Schistosoma japonicum infections were both zero among humans and bovines in 2023, and no S. japonicum infection was detected in snails. These data demonstrated that transmission interruption of schistosomiasis had been achieved across all endemic provinces in China in 2023, and the endemic status of schistosomiasis tended to be stable, while advanced cases were predominant among all schistosomiasis cases. However, the areas of snail habitats remained high and cattle re⁃raising was very common in some regions. Intensified schistosomiasis surveillance and forecast and snail control in high⁃risk areas are needed.

While the priest climbs a post, the devil climbs ten: major biological threats from parasite and vector to malaria control and elimination

YU Xinyu, CAO Jun

2024, 36(3):  228-232,238. 

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Malaria is one of the most serious mosquito⁃borne infectious diseases in the world. The global malaria control progress has stalled in recent years, which is largely due to the biological threats from the malaria pathogen Plasmodium and the vector Anopheles mosquitoes. This article provides an overview of biological threats to global malaria elimination, including antimalarial drug resistance, deletions in the malaria rapid diagnostic test target P. falciparum histidine⁃rich protein 2/3 (Pfhrp2/3) genes, vector insecticide resistance and emergence of invasive vector species, so as to provide insights into malaria and vector research and the formulation and adjustment of the malaria control and elimination strategy.

Biological threats to global malaria eliminationⅠAntimalarial drug resistance

LU Feng

2024, 36(3):  233-238. 

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Malaria is an infectious disease that seriously threatens human health. Currently, malaria control mainly depends on antimalarial chemotherapy. However, antimalarial drug resistance is becoming increasingly severe, which poses a great challenge to malaria control, notably treatment of Plasmodium falciparum malaria. To address this challenge, there is a need to facilitate development of novel antimalarial drugs and innovation of treatment strategies, as well as reinforce surveillance and research on antimalarial drug resistance. This article reviews the main categories and use guidelines of current antimalarial agents, summarizes the current status and monitoring methods of antimalarial drug resistance, and proposes the response to antimalarial drug resistance, so as to provide insights into the use of antimalarial drugs and response to antimalarial drug resistance, and contribute to global malaria elimination.

Biological threats to global malaria eliminationⅡ Deletion in the malaria rapid diagnostic test target Plasmodium falciparum histidine⁃rich protein 2/3 genes

XU Sui, TANG Jianxia

2024, 36(3):  239-242,258. 

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The global malaria epidemic is still severe. Because of simple procedures, rapid detection and accuracy results, rapid diagnostic test (RDT) has become the most important and the most widely used diagnostic tool for malaria prevention and control. However, deletions in the RDT target Plasmodium falciparum histidine⁃rich protein 2/3 (Pfhrp2/3) genes may cause false⁃negative results of RDT, which has been included as one of the four biological threats to global malaria elimination. This article reviews the applications of RDT in the global malaria diagnosis, analyzes the threats and challenges caused by Pfhrp2/3 gene deletion, proposes methods for monitoring Pfhrp2/3 gene deletion, and summarizes the causes and countermeasures of negative RDT detections, so as to provide insights into consolidation of malaria elimination achievements in China and contributions to global malaria elimination.

Biological threats to global malaria elimination Ⅲ Vector insecticide resistance

ZHU Guoding

2024, 36(3):  243-246,271. 

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The insecticide resistance is becoming increasingly severe in malaria vectors and has become one of the most important threats to global malaria elimination. Currently, malaria vectors not only have developed high resistance to conventional insecticides, including organochlorine, organophosphates, carbamates, and pyrethroids, but also have been resistant to recently used neonicotinoids and pyrrole insecticides. This article describes the current status of global insecticide resistance in malaria vectors and global insecticide resistance management strategies, analyzes the possible major challenges in the insecticide resistance management, and proposes the response actions, so as to provide insights into global insecticide resistance management and contributions to global malaria elimination.

Biological threats to global malaria elimination Ⅳ Emergence of invasive vector species

LIU Qiyong

2024, 36(3):  247-250. 

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Driven by international exchanges and climate changes, the invasion and spread of vector Anopheles mosquitoes posed a new challenge to achieving global malaria elimination. Taking the invasion of An. stephensi to exacerbate the malaria epidemic in Africa as an example, this article summarizes the current situation of global Anopheles invasion, and estimates the potential risk of vector Anopheles mosquitoes to unravel the difficulties and challenges in the global malaria elimination program, so as to provide insights into improved early earning and precision control of vector Anopheles mosquito invasion across the world.

Development of a grading diagnostic model for schistosomiasis⁃induced liver fibrosis based on radiomics and clinical laboratory indicators

GUO Zhaoyu, SHAO Juping, ZOU Xiaoqing, ZHAO Qinping, QIAN Peijun, WANG Wenya, HUANG Lulu, XUE Jingbo, XU Jing, YANG Kun, ZHOU Xiaonong, LI Shizhu

2024, 36(3):  251-258. 

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Objective　To investigate the feasibility of developing a grading diagnostic model for schistosomiasis⁃induced liver fibrosis based on B⁃mode ultrasonographic images and clinical laboratory indicators. Methods　Ultrasound images and clinical laboratory testing data were captured from schistosomiasis patients admitted to the Second People’s Hospital of Duchang County, Jiangxi Province from 2018 to 2022. Patients with grade Ⅰ schistosomiasis⁃induced liver fibrosis were enrolled in Group 1, and patients with grade Ⅱ and Ⅲ schistosomiasis⁃induced liver fibrosis were enrolled in Group 2. The machine learning binary classification tasks were created based on patients’ radiomics and clinical laboratory data from 2018 to 2021 as the training set, and patients’ radiomics and clinical laboratory data in 2022 as the validation set. The features of ultrasonographic images were labeled with the ITK⁃SNAP software, and the features of ultrasonographic images were extracted using the Python 3.7 package and PyRadiomics toolkit. The difference in the features of ultrasonographic images was compared between groups with t test or Mann⁃Whitney U test, and the key imaging features were selected with the least absolute shrinkage and selection operator (LASSO) regression algorithm. Four machine learning models were created using the Scikit⁃learn repository, including the support vector machine (SVM), random forest (RF), linear regression (LR) and extreme gradient boosting (XGBoost). The optimal machine learning model was screened with the receiver operating characteristic curve (ROC), and features with the greatest contributions to the differentiation features of ultrasound images in machine learning models with the SHapley Additive exPlanations (SHAP) method. Results　The ultrasonographic imaging data and clinical laboratory testing data from 491 schistosomiasis patients from 2019 to 2022 were included in the study, and a total of 851 radiomics features and 54 clinical laboratory indicators were captured. Following statistical tests (t = -5.98 to 4.80, U = 6 550 to 20 994, all P values < 0.05) and screening of key features with LASSO regression, 44 features or indicators were included for the subsequent modeling. The areas under ROC curve (AUCs) were 0.763 and 0.611 for the training and validation sets of the SVM model based on clinical laboratory indicators, 0.951 and 0.892 for the training and validation sets of the SVM model based on radiomics, and 0.960 and 0.913 for the training and validation sets of the multimodal SVM model. The 10 greatest contributing features or indicators in machine learning models included 2 clinical laboratory indicators and 8 radiomics features. Conclusions　The multimodal machine learning models created based on ultrasound⁃based radiomics and clinical laboratory indicators are feasible for intelligent identification of schistosomiasis⁃induced liver fibrosis, and are effective to improve the classification effect of one⁃class data models. 　

Construction and application of a risk index of Echinococcus infection based on the classification of echinococcosis lesions

XUE Chuizhao, ZHENG Canjun, KUI Yan, SHI Yue, WANG Xu, LIU Baixue, WU Weiping, HAN Shuai

2024, 36(3):  259-271. 

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Objective　To investigate the feasibility of constructing the risk index of Echinococcus infection based on the classification of echinococcosis lesions, so as to provide insights into the management of echinococcosis. Methods　The imaging data of echinococcosis cases were collected from epidemiological surveys of echinococcosis in China from 2012 to 2016, and the detection of incident echinococcosis cases was captured from the annual echinococcosis prevention and control reports across provinces (autonomous regions) and Xinjiang Production and Construction Corps in China from 2017 to 2022. After echinococcosis lesions were classified, a risk index of Echinococcus infection was constructed based on the principle of discrete distribution marginal probability and multi⁃group classification data tests. The correlation between the risk index of Echinococcus infection and the detection of incident echinococcosis cases was evaluated in the provinces (autonomous regions and corps) from 2017 to 2022, and the correlations between the short⁃ and medium⁃term risk indices and between the medium⁃ and long⁃term risk indices of Echinococcus infection were examined using a univariate linear regression model. Results　A total of 4 014 echinococcosis cases in China from 2012 to 2016 were included in this study. The short⁃, medium⁃ and long⁃term risk indices of E. granulosus infection varied in echinococcosis⁃endemic provinces (autonomous regions and corps) of China ([χ2] = 4.12 to 708.65, all P values < 0.05), with high short⁃ (0.058), medium⁃ (0.137) and long⁃term risk indices (0.104) in Tibet Autonomous Region, and the short⁃, medium⁃ and long⁃term risk indices of E. multilocularis infection varied in echinococcosis⁃endemic provinces (autonomous regions and corps) of China ([χ2] = 6.74 to 122.60, all P values < 0.05), with a high short⁃term risk index in Sichuan Province (0.016) and high medium⁃ (0.009) and long⁃term risk indices in Qinghai Province (0.018). There were no significant correlations between the risk index of E. granulosus infection and the detection of incident cystic echinococcosis cases during the study period (t = -0.518 to 2.265, all P values > 0.05), and strong correlations were found between the risk indices of E. multilocularis infection and the detection of incident alveolar echinococcosis cases (including mixed type) in 2018, 2020, 2021, 2022, during the period from 2017 through 2020, from 2017 through 2021, from 2017 through 2022 (all r values > 0.7, t = 2.521 to 3.692, all P values < 0.05). Linear regression models were established between the risk index of E. multilocular infection and the detection of alveolar echinococcosis cases (including mixed type), and the models were all statistically significant (b = 0.214 to 2.168, t = 2.458 to 3.692, F = 6.044 to 13.629, all P values < 0.05). The regression coefficients for the correlations between the medium⁃ and short⁃term, and between the long⁃ and medium⁃term risk indices of E. granulosus infection were 2.339 and 0.765, and the regression coefficients for the correlations between the medium⁃ and short⁃term, and between the long⁃ and medium⁃term risk indices of E. multilocular infection were 0.280 and 1.842, with statistical significance seen in both the regression coefficients and regression models (t = 16.479 to 197.304, F = 271.570 to 38 928.860, all P values < 0.05). Conclusions　The risk index of Echinococcus infection has been successfully established based on the classification of echinococcosis lesions, which may provide insights into the prevention and control, prediction, diagnosis and treatment, and classified management of echinococcosis.

Molecular tracing of Biomphalaria straminea in China

DUAN Lei, QU Lei, GUO Yunhai, GU Wenbiao, LÜ Shan, ZHANG Yi, ZHOU Xiaonong

2024, 36(3):  272-278,285. 

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Objective　To investigate the origin of Biomphalaria straminea in China, so as to provide insights into assessment of schistosomiasis mansoni transmission risk and B. straminea control. Methods　Guanlan River, Dasha River, Shenzhen Reservoir, upper and lower reaches of Kuiyong River, and Xinzhen River in Shenzhen, China, were selected as sampling sites. Ten Biomphalaria samples were collected from each site, and genomic DNA was extracted from Biomphalaria samples. DNA samples were obtained from 15 B. straminea sampled from 5 sampling sites in Minas Gerais State, Pará State, Federal District, Pernambuco State, and Sao Paulo State in Brazil, South America. Cytochrome c oxidaseⅠ(COI) and mitochondrial 16S ribosomal RNA (16S rRNA) genes were sampled using the above DNA templates, and the amplified products were sequenced. The COI and 16S rRNA gene sequences were downloaded from GenBank, and the sampling sites were acquired. All COI and 16S rRNA gene sequences were aligned and evolutionary trees of B. straminea were created based on COI and 16S rRNA gene sequences to identify the genetic similarity and evolutionary relationship between B. straminea samples from China and South America. Results　A total of 60 COI gene sequences with a length of 529 bp and 3 haplotypes were obtained from B. straminea sampled from China. There were 165 COI gene sequences of B. straminea retrieved from GenBank, and following alignment with the above 60 gene sequences, a total of 33 haplotypes were obtained. Phylogenetic analysis showed that the three haplotypes of B. straminea from China were clustered into one clade, among which the haplotype China11 and three B. straminea samples from Brazil retrieved from GenBank belonged to the same haplotype. Geographical evolution analysis showed that the B. straminea samples from three sampling sites along eastern coasts of Brazil had the same haplotype with China11, and B. straminea samples from other two sampling sites were closely, genetically related to China11. A total of 60 16S rDNA gene sequences with approximately 322 bp in length were amplified from B. straminea in China, with 2 haplotypes identified. A total of 70 16S rDNA gene sequences of B. straminea were captured from GenBank. Phylogenetic analysis showed that Biomphalaria snails collected from China were clustered into a clade, and the haplotype China64 and the haplotype 229BS from Brazil shared the same haplotype. The 49 16S rDNA gene sequences of B. straminea from 25 sampling sites in southern Brazil, which were captured from GenBank, were included in the present analysis, and the B. straminea from 3 sampling sites shared the same haplotype with China64 in China. Geographical evolution analysis based on COI and 16S rRNA gene sequences showed that B. straminea sampled from eastern coastal areas of Brazil shared the same haplotypes in two gene fragment sequences with Biomphalaria snails collected from China. Conclusions　The Biomphalaria snails in China are characterized as B. straminea, which have a low genetic diversity. The Biomphalaria snails in China have a high genetic similarity with B. straminea sampled from eastern coastal areas of Brazil, which may have originated from the eastern coastal areas of Brazil.

Preparation and preliminary application of the polyclonal antibody against Toxoplasma gondii dense granule protein 24

FU Shengnan, YANG Yun, WANG Cong, LUO Qingli, YU Li

2024, 36(3):  279-285. 

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Objective　To prepare and characterize the mouse polyclonal antibody against the dense granule protein 24 (GRA24) of Toxoplasma gondii, and explore its preliminary applications. Methods　The GRA24 coding sequences of different T. gondii strains were aligned using the MEGA⁃X software, and the dominant peptide of the GRA24 protein was analyzed with the Protean software. The base sequence encoding this peptide was amplified using PCR assay and ligated into the pET⁃28a vector, and the generated GRA24 truncated protein was transformed into Escherichia coli BL21. After induction by isopropyl⁃beta⁃D⁃thiogalactopyranoside (IPTG), the expression and purification of the recombinant GRA24 protein was analyzed using sodium dodecyl sulfate ⁃ polyacrylamide gel electrophoresis (SDS⁃PAGE). BALB/c mice were immunized by subcutaneous injection with the purified recombinant GRA24 truncated protein to generate the polyclonal antibody, and the titer of the polyclonal antibody was measured using enzyme linked immunosorbent assay (ELISA). The specificity of the polyclonal antibody was tested using Western blotting, and the intracellular localization of the polyclonal antibody was investigated using immunofluorescence assay (IFA). Results　SDS⁃PAGE showed successful construction of the recombinant expression plasmid, and Coomassie brilliant blue staining showed the generation of the high⁃purity recombinant GRA24 truncated protein. ELISA measured that the titer of the polyclonal antibody against the GRA24 truncated protein was higher than 1∶208 400, and Western blotting showed that the polyclonal antibody was effective to recognize the endogenous GRA24 proteins of different T. gondii strains and specifically recognize the recombinant GRA24 truncated protein. Indirect IFA showed that the GRA24 protein secreted 16 hour following T. gondii invasion in host cells. Conclusions　The polyclonal antibody against the T. gondii GRA24 protein has been successfully prepared, which has a widespread applicability, high titers and a high specificity. This polyclonal antibody is available for Western blotting and IFA, which provides the basis for investigating the function of the GRA24 protein.

Effect of oxymatrine on Cryptosporidium parvum infection in mice based on the HMGB1⁃TLR2/TLR4⁃NF⁃κB pathway

SHI Jie, JI Rui, GUAN Zhiyu, ZHANG Xiaoning, LU Yilong

2024, 36(3):  286-293. 

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Objective　To investigate the involvement of the high mobility group box protein B1 (HMGB1)⁃Toll⁃like receptor 2 (TLR2)/TLR4⁃nuclear factor κB (NF⁃κB) pathway in the intestinal mucosal injury induced by Cryptosporidium parvum infection, and to examine the effect of oxymatrine (OMT) on C. parvum infection in mice. Methods　Forty SPF 4⁃week⁃old BALB/c mice were randomly divided into four groups, including the control group, infection group, glycyrrhizin (GA) group and OMT group. Each mouse was orally administered with 1 × 105 C. parvum oocysts one week in the infection, GA and OMT groups following dexamethasone⁃induced immunosuppression to model C. parvum intestinal infections in mice. Upon successful modeling, mice in the GA group were intraperitoneally injected with GA at a daily dose of 25.9 mL/kg for successive two weeks, and animals in the OMT group were orally administered OMT at a daily dose of 50 mg/kg for successive two weeks, while mice in the control group were given normal food and water. All mice were sacrificed two weeks post⁃treatment, and proximal jejunal tissues were sampled. The pathological changes of mouse intestinal mucosal specimens were observed using hematoxylin⁃eosin (HE) staining, and the mouse intestinal villous height, intestinal crypt depth and the ratio of intestinal villous height to intestinal crypt depth were measured. The occludin and zonula occludens protein 1 (ZO1) expression was determined in mouse intestinal epithelial cells using immunohistochemistry, and the relative expression of HMGB1, TLR2, TLR4, myeloid differentiation primary response gene 88 (MyD88) and NF⁃κB p65 mRNA was quantified in mouse jejunal tissues using quantitative real⁃time PCR (qPCR) assay. Results　HE staining showed that the mouse intestinal villi were obviously atrophic, shortened, and detached, and the submucosal layer of the mouse intestine was edematous in the infection group as compared with the control group, while the mouse intestinal villi tended to be structurally intact and neatly arranged in the GA and OMT groups. There were significant differences among the four groups in terms of the mouse intestinal villous height (F = 6.207, P = 0.000 5), intestinal crypt depth (F = 6.903, P = 0.000 3) and the ratio of intestinal villous height to intestinal crypt depth (F = 37.190, P < 0.000 1). The mouse intestinal villous height was lower in the infection group than in the control group ［(321.9 ± 41.1) μm vs. (399.5 ± 30.9) μm; t = 4.178, P < 0.01］ and the GA group ［(321.9 ± 41.1) μm vs. (383.7 ± 42.7); t = 3.130, P < 0.01］, and the mouse intestinal crypt depth was greater in the infection group ［(185.0 ± 35.9) μm］ than in the control group ［(128.4 ± 23.6) μm］ (t = 3.877, P < 0.01) and GA group ［(143.3 ± 24.7) μm］ (t = 2.710, P < 0.05). The mouse intestinal villous height was greater in the OMT group ［(375.3 ± 22.9) μm］ than in the infection group (t = 3.888, P < 0.01), and there was no significant difference in mouse intestinal villous height between the OMT group and the control group (t = 1.989, P > 0.05). The mouse intestinal crypt depth was significantly lower in the OMT group ［(121.5 ± 27.3) μm］ than in the infection group (t = 4.133, P < 0.01), and there was no significant difference in mouse intestinal crypt depth between the OMT group and the control group (t = 0.575, P > 0.05). The ratio of the mouse intestinal villous height to intestinal crypt depth was significantly lower in the infection group (1.8 ± 0.2) than in the control group (3.1 ± 0.3) (t = 10.540, P < 0.01) and the GA group (2.7 ± 0.3) (t = 7.370, P < 0.01), and the ratio of the mouse intestinal villous height to intestinal crypt depth was significantly higher in the OMT group (3.1 ± 0.2) than in the infection group (t = 15.020, P < 0.01); however, there was no significant difference in the ratio of the mouse intestinal villous height to intestinal crypt depth between the OMT group and the control group (t = 0.404, P > 0.05). Immunohistochemical staining showed significant differences among the four groups in terms of occludin (F = 28.031, P < 0.000 1) and ZO1 expression (F = 14.122, P < 0.000 1) in mouse intestinal epithelial cells. The proportion of positive occluding expression was significantly lower in mouse intestinal epithelial cells in the infection group than in the control group ［(14.3 ± 4.5)% vs. (28.3 ± 0.5)%; t = 3.810, P < 0.01］, and the proportions of positive occluding expression were significantly higher in mouse intestinal epithelial cells in the GA group ［(30.3 ± 1.3)%］ and OMT group ［(25.8 ± 1.5)%］ than in the infection group (t = 7.620 and 5.391, both P values < 0.01); however, there was no significant differences in the proportion of positive occluding expression in mouse intestinal epithelial cells between the GA or OMT groups and the control group (t = 1.791 and 2.033, both P values > 0.05). The proportion of positive ZO1 expression was significantly lower in mouse intestinal epithelial cells in the infection group than in the control group ［(14.4 ± 1.8)% vs. (24.2 ± 2.8)%; t = 4.485, P < 0.01］, and the proportions of positive ZO1 expression were significantly higher in mouse intestinal epithelial cells in the GA group ［(24.1 ± 2.3)%］ (t = 5.159, P < 0.01) and OMT group than in the infection group ［(22.5 ± 1.9)%］ (t = 4.441, P < 0.05); however, there were no significant differences in the proportion of positive ZO1 expression in mouse intestinal epithelial cells between the GA or OMT groups and the control group (t = 0.037 and 0.742, both P values > 0.05). qPCR assay showed significant differences among the four groups in terms of HMGB1 (F = 21.980, P < 0.000 1), TLR2 (F = 20.630, P < 0.000 1), TLR4 (F = 17.000, P = 0.000 6), MyD88 (F = 8.907, P = 0.000 5) and NF⁃κB p65 mRNA expression in mouse jejunal tissues (F = 8.889, P = 0.000 7). The relative expression of HMGB1 ［(5.97 ± 1.07) vs. (1.05 ± 0.07); t = 6.482, P < 0.05］、TLR2 ［(5.92 ± 1.29) vs. (1.10 ± 0.14); t = 5.272, P < 0.05］、TLR4 ［(5.96 ± 1.50) vs. (1.02 ± 0.03); t = 4.644, P < 0.05］、MyD88 ［(3.00 ± 1.26) vs. (1.02 ± 0.05); t = 2.734, P < 0.05］ and NF⁃κB p65 mRNA ［(2.33 ± 0.72) vs. (1.04 ± 0.06); t = 2.665, P < 0.05］ was all significantly higher in mouse jejunal tissues in the infection group than in the control group. A significant reduction was detected in the relative expression of HMGB1 (0.63 ± 0.01), TLR2 (0.42 ± 0.10), TLR4 (0.35 ± 0.07), MyD88 (0.70 ± 0.11) and NF⁃κB p65 mRNA (0.75 ± 0.01) in mouse jejunal tissues in the GA group relative to the control group (t = 8.629, 5.830, 11.500, 4.729 and 6.898, all P values < 0.05), and the relative expression of HMGB1, TLR2, TLR4, MyD88 and NF⁃κB p65 mRNA significantly reduced in mouse jejunal tissues in the GA group as compared to the infection group (t = 7.052, 6.035, 4.084, 3.165 and 3.274, all P values < 0.05). In addition, the relative expression of HMGB1 (1.14 ± 0.60), TLR2 (1.00 ± 0.24), TLR4 (1.14 ± 0.07), MyD88 (0.96 ± 0.25) and NF⁃κB p65 mRNA (1.12 ± 0.17) was significantly lower in mouse jejunal tissues in the OMT group than in the infection group (t = 7.059, 5.320, 3.510, 3.466 and 3.273, all P values < 0.05); however, there were no significant differences between the OMT and control groups in terms of relative expression of HMGB1, TLR2, TLR4, MyD88 or NF⁃κB p65 mRNA in mouse jejunal tissues (t = 0.239, 0.518, 1.887, 0.427 and 0.641, all P values > 0.05). Conclusions　C. parvum infection causes intestinal inflammatory responses and destruction of intestinal mucosal barrier through up⁃regulating of the HMGB1⁃TLR2/TLR4⁃NF⁃κB pathway. OMT may suppress the intestinal inflammation and repair the intestinal mucosal barrier through inhibiting the activity of the HMGB1⁃TLR2/TLR4⁃NF⁃κB pathway. 　

Familial aggregation of human hookworm infections in Sichuan Province

LUO Jingwen, TIAN Hongchun, LIU Yang, WU Xiaohong, TIE Lei, ZHANG Liping, DENG Xiu

2024, 36(3):  294-298,328. 

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Objective　To investigate the family aggregation of human hookworm infections in Sichuan Province and to identify its influencing factors, so as to provide insights into management of hookworm infections. Methods　Three to four counties (districts) were sampled from basins, hilly regions and mountainous regions around the basins in Sichuan Province from 2017 to 2022 as fixed survey sites, and 17 to 30 counties (districts) were selected as mobile survey sites. At least 1 000 permanent residents at ages of 3 years and older were sampled from each survey site, and hookworm eggs were detected in human stool samples using the Kato⁃Katz technique. Subjects with 2 and more family members and at least 2 individuals diagnosed with hookworm infections in the county (district) where they lived were selected, and the familial aggregation of hookworm infections was analyzed using the test of goodness of fit for binomial distribution. In addition, the knowledge and practice of hookworm disease control were investigated among residents in Hejiang County and Wutongqiao District, Leshan City, Sichuan Province in 2021 and 2022, and the difference in the knowledge and practice of hookworm disease control was compared between members with and without familial aggregation of hookworm infections. Results　A total of 66 812 residents from 25 196 households were sampled from main endemic areas of hookworm diseases in Sichuan Province from 2017 to 2022 for detection of hookworm infections, and 4 403 infections were identified (6.59% prevalence). The distribution of human hookworm infections in Sichuan Province did not fit the binomial distribution, and showed family aggregations ([χ2] = 2 116.759, P < 0.001). Family aggregation of human hookworm infections was found in endemic areas with 1% and higher prevalence of human hookworm infections ([χ2] = 136.006 to 428.738, all P values < 0.001), and family aggregation of human hookworm infections was identified in different years ([χ2] = 87.615 to 471.838, all P values < 0.001) and in different terrains of endemic areas ([χ2] = 8.423 to 1 144.176, all P values < 0.001). The members with hookworm infections had median eggs per gram of 180 (interquartile range, 780) in aggregated families and 72 (102) in non⁃aggregated families (Z = -2.686, P < 0.05). The proportion of members in families with aggregation of hookworm infections who knew the preventive measures of hookworm disease was significantly lower than in non⁃aggregated families (24.49% vs. 51.72%; [χ2] = 10.262, P < 0.05), and the proportion of members in families with aggregation of hookworm infections who often worked barefoot on the ground was significantly higher than in non⁃aggregated families (30.61% vs. 13.25%; [χ2] = 6.289, P < 0.05). Conclusions There is a familial aggregation of human hookworm infections in Sichuan Province, and awareness of preventive measures for hookworm disease and frequent working barefoot on the ground are associated with familial aggregation of hookworm infections.

Experimental study on the artificial infection of common freshwater snails with Angiostrongylus cantonensis in Dali Bai Autonomous Prefecture, Yunnan Province

LI Tianmei, FANG Wen, CHEN Shaorong, YANG Jing, ZHAO Yongbo, ZHAO Shenhua, LI Ting, YANG Limin, GUO Yunhai, LIU Yuhua

2024, 36(3):  299-303. 

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Objective　To evaluate the potential risk of transmission of angiostrongyliasis by common freshwater snails in Dali Bai Autonomous Prefecture, Yunnan Province, so as to provide insights into local surveillance of angiostrongyliasis. Methods 　 Common freshwater snails were collected from Dali Bai Autonomous Prefecture, Yunnan Province from March to April, 2020, and identified and bred in laboratory. SD rats were infected with third⁃stage larvae of Angiostrongylus cantonensis that were isolated from commercially available Pomacea canaliculata snails in Dali Bai Autonomous Prefecture, and freshwater snails were infected with the first⁃stage larvae of A. cantonensis that were isolated from the feces of SD rats 39 days post⁃infection at room temperature. The developmental process and morphological characteristics of worms in hosts were observed, and the percentages of A. cantonensis infections in different species of freshwater snails were calculated. Then, SD rats were infected with the third⁃stage larvae of A. cantonensis that were isolated from A. cantonensis⁃infected freshwater snails, and the larval development and reproduction was observed. Results　More than 3 000 freshwater snail samples were collected from farmlands, ditches and wetlands around Erhai Lake in Dali Bai Autonomous Prefecture, and Cipangopaludina chinensis, P. canaliculata, Parafossarulus striatulus, Oncomelania hupensis robertsoni, Galba pervia, Physa acuta, Radix swinhoei, Assiminea spp., Tricula spp. and Bellamya spp. were morphologically identified. A total of 105 commercially available P. canaliculata snails were tested for A. cantonensis infections, and 2 P. canaliculata snails were found to be infected with A. cantonensis, in which the third⁃stage larvae of A. cantonensis were isolated. Ten species of freshwater snails were artificially infected with the third⁃stage larvae of A. cantonensis, and all 10 species of freshwater snails were found to be infected with A. cantonensis, with the highest positive rate of A. cantonensis infections in Bellamya spp. (62.3%, 137/204), and the lowest in C. chinensis (35.5%, 11/31). After SD rats were infected with the third⁃stage larvae of A. cantonensis isolated from different species of freshwater snails, mature adult worms of A. cantonensis were yielded. Conclusions　Multiple species of freshwater snails may serve as intermediate hosts of A. cantonensis under laboratory conditions in Dali Bai Autonomous Prefecture of Yunnan Province. Further investigations on natural infection of A. cantonensis in wild snails in Dali Bai Autonomous Prefecture seem justified.

Preliminary observation on the development and dynamic changes of chronic toxoplasmosis in mice

ZHOU Guoqing, BAI Shaoyuan, LI Yuyuan, ZHU Guoding, HUANG Siyang

2024, 36(3):  304-309. 

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Objective　To investigate the development and dynamic changes of cysts in the brain of mice following infection with different forms of Toxoplasma gondii, so as to provide insights into for toxoplasmosis prevention and control. Methods　ICR mice at ages of 6 to 8 weeks, each weighing 20 to 25 g, were intraperitoneally injected with tachyzoites of the T. gondii PRU strain at a dose of 1 × 105 tachyzoites per mouse, orally administered with cysts at a dose of 20 oocysts per mouse or oocysts at a dose of 200 oocysts per mouse for modeling chronic T. gondii infection in mice, and the clinical symptoms and survival of mice were observed post⁃infection. Mice were orally infected with T. gondii cysts at doses of 10 (low⁃dose group), 20 (medium⁃dose group), 40 cysts per mouse (high⁃dose group), and the effect of different doses of T. gondii infections on the number of cysts was examined in the mouse brain. Mice were orally administered with T. gondii cysts at a dose of 20 cysts per mouse, and grouped according to gender (female and male) and time points of infections (20, 30, 60, 90, 120, 150, 180 days post⁃infection), and the effects of gender and time points of infections on the number of cysts was examined in the mouse brain. In addition, mice were divided into the tachyzoite group (Group T), the first⁃generation cyst group (Group C1), the second⁃generation cyst group (Group C2), the third⁃generation cyst (Group C3) and the fourth⁃generation cyst group (Group C4). Mice in the Group T were intraperitoneally injected with T. gondii tachyzoites at a dose of 1 × 105 tachyzoites per mouse, and the cysts were collected from the mouse brain tissues 30 days post⁃infection, while mice in the Group C1 were orally infected with the collected cysts at a dose of 30 cysts per mouse. Continuous passage was performed by oral administration with cysts produced by the previous generation in mice, and the effect of continuous passage on the number of cysts was examined in the mouse brain. Results　Following infection with T. gondii tachyzoites, cysts and oocysts in mice, obvious clinical symptoms were observed on days 6 to 113 and mice frequently died on days 7 to 12. The survival rates of mice were 67.0%, 87.0% and 53.0%, and the mean numbers of cysts were (516.0 ± 257.2), (1 203.0 ± 502.0) and (581.0 ± 183.1) in the mouse brain (F = 11.94, P < 0.01) on day 30 post⁃infection with T. gondii tachyzoites, cysts and oocysts, respectively, and the numbers of cysts in the brain tissues were significantly lower in mice infected with T. gondii tachyzoites and oocysts than in those infected with cysts (all P values < 0.01). The survival rates of mice were 87.0%, 87.0% and 60.0%, and the mean numbers of cysts were (953.0 ± 355.5), (1 084.0 ± 474.3) and (1 113.0 ± 546.0) in the mouse brain in the low⁃, medium⁃ and high⁃dose groups on day 30 post⁃infection, respectively (F = 0.42, P > 0.05). The survival rates of male and female mice were 73.0% and 80.0%, and the mean numbers of cysts were (946.4 ± 411.4) and (932.1 ± 322.4) in the brain tissues of male and female mice, respectively (F = 1.63, P > 0.05). Following continuous passage, the mean numbers of cysts were (516.0 ± 257.2), (1 203.0 ± 502.0), (896.8 ± 332.3), (782.5 ± 423.9) and (829.2 ± 306.0) in the brain tissues of mice in the T, C1, C2, C3 and C4 groups, respectively (F = 4.82, P < 0.01), and the number of cysts was higher in the mouse brain in Group 1 than in Group T (P < 0.01). Following oral administration of 20 T. gondii cysts in mice, cysts were found in the moues brain for the first time on day 20 post⁃infection, and the number of cysts gradually increased over time, peaked on days 30 and 90 post⁃infection and then gradually decreased; however, the cysts were still found in the mouse brain on day 180 post⁃infection. Conclusions　There is a higher possibility of developing chronic T. gondii infection in mice following infection with cysts than with oocysts or tachyzoites and the most severe chronic infection is seen following infection with cysts. The number of cysts does not correlate with the severity of chronic T. gondii infection, and the number of cysts peaks in the mouse brain on days 30 and 90 post⁃infection.

Auxiliary diagnostic value of T cells spot test of Mycobacterium tuberculosis infection for pulmonary and extra⁃pulmonary tuberculosis among the elderly

HUANG Rui, LI Shuai, WANG Changmin

2024, 36(3):  310-313. 

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Objective　To evaluate the auxiliary diagnostic value of T cells spot test of Mycobacterium tuberculosis infection (T⁃SPOT.TB) for pulmonary and extra⁃pulmonary tuberculosis among the elderly. Methods　A total of 173 elderly patients at ages of 60 years and older and with suspected tuberculosis that were admitted to People’s Hospital of Xinjiang Uygur Autonomous Region during the period from October 2022 through February 2024 were enrolled, and all patients underwent T⁃SPOT.TB, acid fast staining and GeneXpert MTB/RIF tests. The etiological tests of MTB served as a gold standard, and the diagnostic values of T⁃SPOT.TB, acid fast staining and GeneXpert MTB/RIF tests for pulmonary and extra⁃pulmonary tuberculosis were compared among the elderly patients. Results　Of the 173 elderly patients suspected of tuberculosis, there were 44 patients definitely diagnosed with pulmonary tuberculosis, 30 cases with extra⁃pulmonary tuberculosis, and 99 cases without tuberculosis. The sensitivities of T⁃SPOT.TB, acid fast staining and GeneXpert MTB/RIF tests were 86.5%, 27.0% and 54.1% for diagnosis of tuberculosis. The sensitivities of T⁃SPOT.TB were 86.4% and 86.7% for diagnosis of pulmonary tuberculosis and extra⁃pulmonary tuberculosis, with an 80.8% specificity for diagnosis of tuberculosis. The sensitivities of GeneXpert MTB/RIF were 56.8% and 50.0% for diagnosis of pulmonary tuberculosis and extra⁃pulmonary tuberculosis, with a 100.0% specificity each, and the sensitivities of acid fast staining were 31.8% and 20.0% for diagnosis of pulmonary tuberculosis and extra⁃pulmonary tuberculosis, with a 100.0% specificity each. In addition, the areas under the receiver operating characteristic curve were 0.836, 0.635 and 0.770 for diagnosis of tuberculosis with T⁃SPOT.TB, acid fast staining and GeneXpert MTB/RIF tests among the elderly patients, respectively. Conclusion　T⁃SPOT.TB has a high auxiliary diagnostic value for both pulmonary and extra⁃pulmonary tuberculosis among elderly patients.

Application of the CRISPR/Cas system in gene editing and nucleic acid detection of parasitic diseases: a review

YAN Shuning, YANG Shuo, YANG Hanyin, XIN Yi, XU Bin, HU Wei, LU Yan, ZHENG Bin

2024, 36(3):  314-320. 

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CRISPR/Cas system, an adaptive immune system with clustered regularly interspaced short palindromic repeats, may interfere with exogenous nucleic acids and protect prokaryotes from external damages, is an effective gene editing and nucleic acid detection tools. The CRISPR/Cas system has been widely applied in virology and bacteriology; however, there is relatively less knowledge about the application of the CRISPR/Cas system in parasitic diseases. The review summarizes the mechanisms of action of the CRISPR/Cas system and provides a comprehensive overview of their application in gene editing and nucleic acid detection of parasitic diseases, so as to provide insights into future studies on parasitic diseases.

How do female mosquitoes determine the most suitable males for mating?

LI Yitong, LI Dong, LIU Xiaofei, WANG Ying, LIU Tingting, XU Yanqiu, DENG Shengqun

2024, 36(3):  321-328. 

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More than 80% of the world’s populations are at risk of vector⁃borne diseases, with mosquito⁃borne diseases as a significant global public health problem. Mosquito populations control is critical to interrupting the transmission of mosquito⁃borne diseases. This review summarizes the physical attributes, smell, vision, touch, and hearing of mosquitoes to unravel the preferences of female mosquitoes, and describes the mechanisms underlying the best male mating by female mosquitoes, so as to provide new insights into management of mosquito⁃borne diseases.