Table of Content

30 October 2024, Volume 42 Issue 5

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SPECIAL REPORT

Prevalence of visceral leishmaniasis in China in 2023

ZHOU Zhengbin, LI Yuanyuan, LI Zhongqiu, YANG Limin, LIU Qin, ZHANG Yi, CHEN Junhu, LI Shizhu

2024, 42(5):  559-565.  doi:10.12140/j.issn.1000-7423.2024.05.001

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Objective To understand the epidemic situation of visceral leishmaniasis (VL) in China in 2023 to provide scientific evidence for formulating prevention and control strategies. Methods Data on VL cases reported in 2023 was collected from the web-based National Diseases Reporting Information System (NDRIS) operated by the Chinese Center for Disease Control and Prevention. From the collected information, suspected cases, duplicate cases and cutaneous leishmaniasis cases were excluded to establish a database. Descriptive epidemiological analysis of the three-dimensional distribution of VL was conducted using Microsoft Excel 2016 software. Results A total of 299 VL cases were reported in 125 counties across 15 provinces (autonomous regions and municipalities) in 2023, with an incidence rate of 0.02/100 000, showing a 25.1% increase from the year 2022. Among these, 237 cases were of mountain-type zoonotic VL (MT-VL), 7 cases were of desert-type zoonotic VL (DT-ZVL), 1 case was of anthroponotic VL (AVL), and 54 cases were imported cases from non-endemic regions. The reported cases were mainly distributed in Shanxi (114 cases), Henan (54 cases) and Shaanxi (38 cases) provinces, together accounting for 68.9% (206/299) of the total reported cases in China. Of the 125 counties, 74 were endemic counties, reporting 245 locally acquired cases, while the remaining 51 were non-endemic counties, reporting 54 imported cases. Among the 74 endemic provinces, seven counties including Pingding County, Suburb District of Yangquan, Xiangfen County in Shanxi Province, Jingxing County of Hebei Province, Huazhou District in Shaanxi Province, Linzhou City and Xin’an County of Henan Province were the major endemic counties. These counties reported 24, 16, 10, 13, 12, 12 and 10 VL cases, respectively, accounting for 32.4% (97/299) of the total cases reported nationwide. Eighteen counties, including Hongdong, Licheng, Linxian, Shilou, and Wutai counties in Shanxi Province; Quyang, Fuping, Laiyuan, Pingshan, Shexian, Tangxian, and Yixian counties in Hebei Province; Songxian and Yuzhou in Henan Province; Chengxian and Kangxian counties in Gansu Province; Yanqing District in Beijing; and Chengcheng County in Shaanxi Province, were new reemergence VL endemic counties, reporting 23 locally acquired cases. The peak incidence of VL occurred in March. The ratio of male to female cases was 1:0.4. Farmers accounted for 57.2% (171/299) of the total cases. The age distribution of VL cases was predominantly in the 45-74 years old group (64.5%). Conclusion VL in China exhibits a low prevalence but the incidence showed a rapid increasing trend, and the endemic area was in expanding. Farmers are recognized as the high-risk population for visceral leishmaniasis. It is imperative to strengthen the surveillance and prevention of visceral leishmaniasis.

ORIGINAL ARTICLES

Study on mechanism of Echinococcus granulosus antigen B and calcium binding protein 1 regulating immune thrombocytopenia in mice

YANG Xuehua, SONG Haichen, JIAO Hongjie, CHENG Yongfeng, YUE Yingbin, SONG Chuanlong, HE Baiqifeng, YAN Mei

2024, 42(5):  566-572.  doi:10.12140/j.issn.1000-7423.2024.05.002

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Objective To investigate the protective effects and mechanisms of Echinococcus granulosus antigen B (AgB) and calcium-binding protein 1 (CBP1) in immune thrombocytopenia (ITP) mouse models through TLR4/NF-κB/NLRP3 signaling pathway. Methods Forty-two healthy female BALB/c mice were randomly divided into six groups: control, ITP, AgB, AgB + ITP, CBP1 and CBP1 + ITP, with seven mice in each group. Mice in the AgB and AgB + ITP groups received daily intraperitoneal injection of 100 μg (200 μl) of AgB, while those in the CBP1 and CBP1 + ITP groups received 100 μg (200 μl) of CBP1, mice in the control and ITP groups received 200 μl of PBS, all for five consecutive days. Subsequently, mice in the ITP, AgB + ITP and CBP1 + ITP groups received daily intraperitoneal injection of 3 μg (200 μl) of anti-CD41 monoclonal antibody (Ab) to establish ITP model, while those in the control, AgB, and CBP1 groups received 200 μl of PBS daily, all for five consecutive days. Platelet counts in peripheral blood were measured one day before modeling (D0), during modeling (D1-D5), and one day after modeling (D6). On the day after modeling, the mice were euthanized to collect spleens and livers, which were weighed for calculation of organ index. The mRNA relative transcription levels of Toll-like receptor 4 (TLR4), NOD-like receptor pyrin domain-containing protein 3 (NLRP3), apoptosis-associated speck-like protein containing a CARD (ASC) and IL-1β in spleen tissues were detected by qRT-PCR. The relative expression levels of TLR4, inducible nitric oxide synthase (iNOS), nuclear factor κB (NF-κB), and caspase-1 in spleen tissues were detected by Western blotting. One-way ANOVA was used for comparisons between groups, and the LSD-t test was used for pairwise comparison between multiple groups. Results From the D1 to D6, the platelet counts in the ITP group mice were (102.1 ± 6.8) × 10⁹/L, (234.7 ± 18.1) × 10⁹/L, (229.7 ± 45.8) × 10⁹/L, (316.7 ± 26.8) × 10⁹/L, (320.6 ± 60.5) × 10⁹/L, (179.1 ± 22.2) × 10⁹/L, which were lower than those of (526.6 ± 90.4) × 10⁹/L, (679.3 ± 58.5) × 10⁹/L, (828.0 ± 61.0) × 10⁹/L, (855.3 ± 101.9) × 10⁹/L, (784.1 ± 177.7) × 10⁹/L, (877.4 ± 107.5) × 10⁹/L in the control group (LSD-t = -4.2, -5.5, -6.9, -6.3, -3.9, -4.8; all P < 0.05). Platelet counts in the mice of AgB + ITP group and CBP1 + ITP group on D6 were (512.6 ± 100.5) × 10⁹/L and (511.1 ± 114.8) × 10⁹/L, which were higher than those in the ITP group (LSD-t = 2.3, 2.3; both P < 0.05). The spleen index of the ITP group was 12.1 ± 1.2, higher than that of the control group (6.3 ± 0.4) (LSD-t = 6.8, P < 0.01). The spleen index in the AgB + ITP group was 9.0 ± 0.3, which was lower than that in the ITP group (LSD-t = -3.6, P < 0.01). The results of qRT-PCR showed that the relative transcription levels of TLR4, NLRP3, ASC, IL-1β in spleen tissue of the ITP group were 7.5 ± 2.1, 5.3 ± 1.5, 3.6 ± 0.7, 4.0 ± 0.9, respectively, which were higher than those of 1.3 ± 0.3, 1.2 ± 0.2, 1.2 ± 0.3, 1.0 ± 0.1 in the control group (LSD-t = 4.8, 4.5, 4.2, 5.2; P < 0.01). The mRNA relative transcription levels of the AgB + ITP group were 1.7 ± 0.3, 0.6 ± 0.1, 1.0 ± 0.3 and 0.7 ± 0.1, while those of CBP1 + ITP group were 1.7 ± 0.1, 1.0 ± 0.3, 1.1 ± 0.4 and 0.4 ± 0.1, all were lower than those of ITP group (LSD-t = -4.6, -5.1, -4.5, -5.9, -4.5, -4.7, -4.4, -6.3; all P < 0.01). Western blotting results showed that the relative expression levels of TLR4, iNOS, NF-κB p65, caspase-1 in spleen tissue of the ITP group were 0.7 ± 0.1, 0.5 ± 0.0, 1.4 ± 0.2, 1.5 ± 0.2, all were higher than those of 0.2 ± 0.0, 0.3 ± 0.0, 0.9 ± 0.2, 0.8 ± 0.2 in the control group (LSD-t = 8.6, 6.5, 3.1, 3.5; all P < 0.01). The relative expression levels of AgB + ITP group were 0.4 ± 0.0, 0.4 ± 0.0, 0.9 ± 0.1, 1.0 ± 0.1, while those of CBP1 + ITP group were 0.2 ± 0.1, 0.2 ± 0.0, 0.6 ± 0.1, 0.7 ± 0.1, all were lower than those of ITP group (LSD-t = -6.1, -2.8, -3.1, -2.3, -8.9, -7.1, -4.9, -4.2; all P < 0.05). Conclusion Both AgB and CBP1 showed protective effect in mouse ITP, and their mechanisms are associated with the regulation of the TLR4/NF-κB/NLRP3 pathway.

Treatment effect of Zhuang medicine Herba abri formula on hepatic fibrosis induced by Echinococcus multilocularis infection

LI Jiajing, HUANG Wenjun, CAO Deping

2024, 42(5):  573-581.  doi:10.12140/j.issn.1000-7423.2024.05.003

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Objective To observe the treatment effect of Zhuang medicine Herba abri formula (HaF) on hepatic fibrosis induced by Echinococcus multilocularis infection in vivo and in vitro. Methods The HaF solution was prepared from Zhuang medicine Herba abri capsule commercial available. Human hepatic stellate cell-LX2 (HSC-LX2) were cultured with 25, 50, 100 or 200 µg/ml HaF solution for 72, 96 and 120 h, then the relative expression of α-smooth muscle actin (α-SMA) and collagen Ⅰ in HSC-LX2 was detected with Western blotting, respectively. Separately, HSC-LX2 cells were stimulated with 0.1 mg/ml E. multilocularis crude antigen for 24 h, followed by addition of 100 μg/ml HaF for 24, 48 and 72 h, subsequently, the relative expression of α-smooth muscle actin (α-SMA) and collagen Ⅰ in HSC-LX2 were detected by Western blotting. Thirty-six female Kunming mice were randomly assigned into the following groups: control group (4 mice), infection group (4 mice), HaF treatment groups at doses of 0.5 g/(kg•d) (5 mice), 1.0 g/(kg•d) (5 mice), and 2.0 g/(kg•d) (6 mice), albendazole (ABZ) treatment group (6 mice), and ABZ + HaF combined treatment group (6 mice). The infection group and each treatment group received intraperitoneal injection of 2 000 protoscolices, while the control group was injected with an equal volume of normal saline. After a period of 60 days post-infection, the mice in treatment groups were treated daily by gavage with the corresponding medication for 60 days, and then the mice liver and spleen tissues were collected and weighed to calculate hepatic index and splenic indix. The content of hydroxyproline (HYP) in liver tissue was determined by the alkaline hydrolysis method, and the level of serum aspartate aminotransferase (AST) was measured by the colorimetric method. Western blotting was performed to detect the relative expression of α-SMA and collagen Ⅰ in the liver, while hematoxylin-eosin staining (HE) and Masson staining were used to observe pathological changes in liver and spleen tissues. The comparison between two groups of data was conducted using an independent-samples t-test. Results The relative expression of α-SMA protein and collagen Ⅰ was 0.401 ± 0.218 and 0.352 ± 0.058, respectively, in HSC-LX2 cells treated with 100 µg/ml HaF solution at 96 h, which were significantly lower than those in the control group (1.435 ± 0.297, 1.340 ± 0.416) (t = 2.755, 11.120; both P < 0.05). The control group did not exhibit any statistically significant differences compared to the other concentrations and durations of treatment. Compared to the control group (0.895 ± 0.417, 1.009 ± 0.378), the relative expression levels of α-SMA and collagen Ⅰ in HSC-LX2 treated with HaF for 48 h after E. multilocularis crude antigen stimulation significantly decreased to (0.326 ± 0.106) and (0.315 ± 0.076), respectively (t = 6.359, 9.059; both P < 0.05). In vivo experiments showed that the hepatic indices, splenic indices, serum AST levels and liver HYP contents of infection group were (4.366 ± 0.284) %, (5.129 ± 1.114) mg/g, (22.194 ± 1.509) U/L, and (21.743 ± 2.503) × 10^-2 μg/mg respectively, which were higher than (3.389 ± 0.045) %, (2.031 ± 0.165) mg/g, (17.355 ± 0.574) U/L and (9.330 ± 1.519) × 10^-2 μg/mg in the control group (t = 3.393, 2.752, 2.616, 4.549; all P < 0.05). The serum AST levels of mice in the HaF treatment group at doses of 0.5, 1.0, and 2.0 g/(kg·d), the ABZ treatment group, and the combined ABZ + HaF treatment group were (17.375 ± 0.746), (15.411 ± 1.338), (17.057 ± 1.066), (16.190 ± 1.559), (14.637 ± 1.888) U/L, respectively, lower than those in infection group (t = 2.863, 3.362, 2.838, 2.717, 3.002; all P < 0.05). Among the treatment groups, the HaF treatment group at a dosage of 1.0 g/(kg•d) showed the most significant reduction in serum AST levels. There were no significant differences in hepatic/splenic indices and HYP content in liver tissue between the five treatment groups and the infection group (all P > 0.05). Western blotting results showed that the relative expression level of α-SMA protein in the liver tissue of mice in the 1.0 g/(kg•d) HaF treatment group was 0.818 ± 0.195, and the relative expression level of collagen Ⅰ in the liver tissue of mice in the combined treatment group of ABZ and HaF was 0.799 ± 0.167, both lower than those in the infected group (1.668 ± 0.131, 1.341 ± 0.141) (t = 3.619, 2.486; P < 0.05). The results of HE staining and Masson staining demonstrated that the fibrotic tissue area in the ABZ + HaF combined treatment group was (6.184 ± 1.951)%, which was significantly reduced compared to (19.916 ± 2.046)% in the infection group (t = 4.703, P < 0.01), indicating a notable improvement in liver fibrosis. Conclusion Zhuang medicine HaF demonstrates the ability to attenuate hepatic fibrosis induced by E. multilocularis infection both in vitro and in vivo. It also provides an experimental basis for HaF as a novel therapeutic agent for treatment of alveolar echinococcosis.

Identification of lesion activities in haptic cystic echinococcosis using machine learning model based on radiomics and clinical features

WANG Zhanjin, CHEN Zhiheng, LI Fuyuan, CAI Junjie, XUE Zhangtuo, ZHOU Ying, CAO Yuntai, WANG Zhan

2024, 42(5):  582-593.  doi:10.12140/j.issn.1000-7423.2024.05.004

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Objective To develop machine learning models utilizing radiomic and clinical features to precisely identify the biological activity of haptic cystic echinococcosis (HCE). Methods The CT images and clinical data of 521 HCE patients treated at the Hepatobiliary and Pancreatic Surgery Department of Qinghai University Affiliated Hospital, along with 236 HCE patients treated at the General Surgery Departments of Guoluo Prefectural People’s Hospital and Yushu Prefectural People’s Hospital in 2018-2022, were collected. Radiomics features were extracted and screened accordingly. Univariate and multivariate logistic regression analyses were performed on the clinical data to select features for model construction. To construct radiomics and clinical models, seven machine learning algorithms were employed including Logistic Regression (LR), Support Vector Machine (SVM), K-Nearest Neighbors (KNN), Random Forest (RandomForest), Extreme Gradient Boosting (XGBoost), Light Gradient Boosting Machine (LightGBM), and Extra Trees. A clinical-image combined model was constructed based on the prediction from radiomics model combining clinical model, using soft voting method. DeLong’s test was used to compare the performances of the radiomics model, clinical model, and combined clinical-imaging model. In addition, external validation was utilized to assess the model’s performance. Results A total of 430 patients were included for model development and training, while 171 patients were designated for external validation. Fifty-one radiomics features and five clinical features were selected for model construction. Among the seven machine learning models, the XGBoost algorithm demonstrated the best performance, achieving area under the curve (AUC) values of 0.977 [95% confidence interval (CI): 0.964-0.990] and 0.839 (95% CI: 0.776-0.901) on the training and external validation sets, respectively. The radiomics model achieved AUC values of 0.998 (95% CI: 0.997-1.000) and 0.874 (95% CI: 0.822-0.927), while the combined model obtained AUC values of 1.000 (95% CI: 0.999-1.000) and 0.931 (95% CI: 0.894-0.968). The DeLong test results indicated that the performance of the combined model was superior to that of the clinical model in the training set (Z = 2.154, P < 0.05) and showed no statistically significant difference when compared to the radiomics model (Z = 0.562, P > 0.05); however, its performance on the external validation set was better than both the clinical and radiomics models (Z = 3.338, 3.331; P < 0.05). Calibration plots and decision curve analysis (DCA) indicated that the combined model exhibited the best calibration performance in both the training and external validation sets, yielding the highest net benefit, demonstrating consistent performance across different datasets, and displaying good generalizability and reliability in external validation. Conclusion The machine learning model, developed based on radiomic and clinical data, can precisely identify the biological activity of HCE lesions. The combined model exhibits higher diagnostic accuracy and clinical application potential, providing reference for making treatment plan for HCE patients.

Effect of NK1.1 expression on liver T cell function in the advanced stage of mice infected with Echinococcus multilocularis

ZHAO Hanyue, LI Jintian, XUE Junlong, KALIBIXIATI Aimulajiang, LIN Renyong, TUERGANAILI Aji

2024, 42(5):  594-600.  doi:10.12140/j.issn.1000-7423.2024.05.005

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Objective To observe the changes in NK1.1 expression of liver T cells in the advanced stage of mice infected with Echinococcus multilocularis, and analyze the effect of NK1.1 expression on liver T cells’ function. Methods C57BL/6N mice were assigned into the infection group and the control group randomly. Mice in the infection group were injected with 2 000 E. multilocularis protoscoleces through hepatic portal vein, while mice in the control group injected with the same volume of normal saline. The livers were collected at the advanced stage of infection (36 weeks post-infection). The pathological changes of the liver tissues were observed by hematoxylin-eosin (HE) staining, and the degree of live fibrosis was observed by Masson staining and Sirius red staining. The lymphocytes were collected and the flow cytometry was performed to detect the proportions of CD4⁺ T and CD8⁺ T cells. The expression changes of NK1.1, granzyme B (GrB), perforin, interferon-γ (IFN-γ) and interleukin-17 (IL-17) in different T cells were also detected. SPSS 25.0 software was used for statistical analysis, and an independent sample t-test was used to compare the mean between the two groups. Results HE staining showed that lesions with germinal layer structure were formed in the livers of mice in the infection group 36 weeks post-infection, with infiltration of inflammatory cells in surrounding tissues, and results of Masson staining and Sirius red staining showed the significant hepatic fibrosis. Flow cytometry results showed the proportions of CD4⁺ T and CD8⁺ T cells in the liver of mice in the infection group 36 weeks post-infection were (32.0 ± 6.0)% and (42.0 ± 13.9)%, respectively, which were (36.6 ± 9.0)% and (50.2 ± 5.2)% in the control group, with no significant differences (t = 1.068, 1.446, both P > 0.05). The proportion of NK1.1⁺CD4⁺ T cells in the infection group was (30.0 ± 14.5)%, which was lower than NK1.1⁺CD4⁺ T cells in the control group [(69.0 ± 10.7)%] (t = 5.573, P < 0.01) and NK1.1^-CD4⁺ T cells in the infection group [(51.6 ± 10.8)%] (t = 2.920, P < 0.05). The proportion of NK1.1⁺CD8⁺ T cells in the infection group was (22.0 ± 12.9)%, which was lower than NK1.1⁺CD8⁺ T cells in the control group [(43.6 ± 5.9)%] (t = 3.999, P < 0.01) and NK1.1^-CD8⁺ T cells in the infection group [(64.0 ± 13.6)%] (t = 5.496，P < 0.01). The proportions of NK1.1⁺CD4⁺ T cells that secreted GrB, IFN-γ and IL-17 in the infection group were (4.8 ± 2.8)%, (6.8 ± 6.0)% and (6.9 ± 5.2)%, respectively, which were lower than those in the control group [(12.6 ± 4.8)%, (23.6 ± 7.3)% and (22.1 ± 7.1)%] (t = 3.462, 4.171, 4.124, all P < 0.01). The proportions of NK1.1⁺CD8⁺ T cells that secreted GrB, IFN-γ and IL-17 in the infection group were (1.5 ± 0.9)%, (5.3 ± 2.7)% and (3.7 ± 2.3)%, respectively, which were lower than those in the control group [(6.0 ± 2.7)%, (18.1 ± 9.5)% and (16.9 ± 9.3)%] (t = 3.861, 3.196, 3.361, P < 0.01, 0.05, 0.01). The proportions of NK1.1⁺CD4⁺ T and NK1.1⁺CD8⁺ T cells that secreted perforin in the infection group were (3.0 ± 1.3)% and (2.7 ± 1.2)%, respectively, which were (2.3 ± 1.2)% and (2.1 ± 1.6)% in the control group, with no significant differences (t = 1.027, 0.717, both P > 0.05). In the infection group, the proportion of NK1.1^-CD4⁺ T cells secreted GrB was (6.7 ± 2.1)%, which was higher than NK1.1⁺CD4⁺ T cells (t = 2.629, P < 0.05); the proportion of NK1.1^-CD8⁺ T cells secreted IFN-γ was (13.7 ± 9.2)%, which was higher than NK1.1⁺CD8⁺ T cells (t = 2.609, P < 0.05); the proportion of NK1.1^-CD8⁺ T cells secreted IL-17 was (1.0 ± 0.4)%, which was lower than NK1.1⁺CD8⁺ T cells (t = 2.740, P < 0.05). Conclusion The number of NK1.1⁺CD4⁺ T and NK1.1⁺CD8⁺ T cells decreased in the liver of mice infected with E. multilocularis in the advanced stage, and the secretion of GrB, IFN-γ and IL-17 were weakened, which indicated the existence of immune depletion of NK1.1⁺ T cells.

Analysis of surveillance results on schistosomiasis prevalence post-reassessment of elimination in five provinces under schistosomiasis elimination program

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

2024, 42(5):  601-607.  doi:10.12140/j.issn.1000-7423.2024.05.006

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Objective To analyze the schistosomiasis epidemic situation post-reassessment in five provinces (Municipality, Autonomous Region) that have achieved schistosomiasis elimination as of the end of 2023, including Shanghai, Zhejiang, Fujian, Guangdong, and Guangxi, and to provide scientific evidence for further control and elimination of schistosomiasis. Methods The data relevant to schistosomiasis control and including examinations in residents and livestock, snail survey and molluscaciding in endemic counties, cities, and districts of Shanghai, Zhejiang, Fujian, Guangdong, and Guangxi in 2015 to 2023 were collected from National Schistosomiasis Control and Prevention Information Management System, as well as schistosomiasis case report information from China CDC management System of Systematic Surveillance Reports of Disease Prevention and Control Information. The data collected were analyzed. Results From 2015 to 2023, 64 cases of schistosomiasis were reported in five provinces, including 50 cases of schistosomiasis japonica imported from other provinces in China and 14 cases of African schistosomiasis imported from other countries. Schistosomiasis eleimination achieved in 112 historically endemic counties in the five provinces, where the number of schistosomiasis cases decreased from 1 038 in 2015 to 780 in 2023, showing a 24.86% reduction, and no local infection cases were reported. The existing schistosomiasis cases were comprised of local advanced schistosomiasis (1 028 cases) and imported schistosomiasis cases (10 cases), all of which were distributed in Zhejiang Province. The overall serological tests positive rate was 0.59% in 2015 (827/141 204) to 0.17% in 2023 (228/136 002). Among them, the average sero-positive rate in Changshan County of Zhejiang Province was the highest (1.87%, 1 469/78 484). The number of bovines decreased by 51.93% from 125 362 in 2015 to 60 267 in 2023, 29 298 received serological examinations, with 8 positive detected in 2015 only. A total of 4 284 bovines received stool examinations, with no positives identified. From 2015 to 2023, snail distribution was reported in 38 counties. The actual snail habitats increased from 75.26 hm² (1 hm² = 10 000 m²) in 2015 to 86.35 hm² in 2023. Hill and water network types were the main snail habitat types. From 2015 to 2018, the area of emerging snail habitats increased annually, peaking at 12.57 hm² in 2018, while from 2019 to 2023, the area was fluctuated and decreased to 0.56 hm² by 2023. From 2015 to 2023, molluscicide treatment was annually performed in 2 076.95 hm² to 2 307.35 hm² snail habitats, 19 735.08 hm² snail habitats received molluscicide treatment in total throughout the 9 years. Environmental improvements were performed in snail habitats covering an area of 110.63 hm², among which Guangdong Province had the largest improved area (58.53 hm², 52.91%). Conclusion The five provinces have been continueing to consolidate the achievements of schistosomiasis elimination. It is recommended that the five provinces persist the control concept of surveying and eliminating remaining snails and preventing exogenous sources of infection, underline the issue of emerging and reemerging snail habitats, and continuously strengthen risk surveillance and epidemic information management to consolidate the achievements in schistosomiasis elimination.

Establishment and application evaluation of a rapid visualization detection method for Schistosoma japonicum specific nucleic acid fragments based on RPA-CRISPR/Cas12a technology

XU Guolei, FENG Yanye, HU Wei

2024, 42(5):  608-614.  doi:10.12140/j.issn.1000-7423.2024.05.007

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Objective To develop a rapid visualization method for specific nucleic acid fragments of Schistosoma japonicum using the recombinase polymerase amplification (RPA) technology combined with clustered regularly interspaced short palindromic repeats-associated 12a protein (CRISPR/Cas12a), and to preliminarily evaluate its application value. Methods RPA primers, single-stranded DNA (ssDNA) reporter and CRISPR RNA (crRNA) were designed to target the SjCHGCS20 sequence (GenBank: FN356222.1). The concentrations of T4 recombinase Y, T4 recombinase X, T4 single-stranded binding protein and creatine kinase were optimized in the RPA reaction system, along with the RPA reaction temperature and time. A one-tube RPA-CRISPR/Cas12a method was established by adding the optimized RPA amplification assays, mineral oil and Cas12a reaction system to a reaction tube. After RPA amplification under optimized conditions, the Cas12a reaction was initiated by mixing with the RPA amplification products. The bright green fluorescence signal, indicative of a positive result, was visualized following the CRISPR/Cas12a-mediated cleavage reaction. The sensitivity of the RPA-CRISPR/Cas12a method was evaluated by detecting various amounts of S. japonicum genomic DNA (10⁰, 10^-1, 10^-2, 10^-3, 10^-4, 10^-5 and 10^-6 ng). The assay specificity was assessed by detecting genomic DNA (1 ng total) from S. japonicum, S. mansoni, Paragonimus westermani and Clonorchis sinensis. The effectiveness of the method was further evaluated using mice dummy positive sera containing S. japonicum genomic DNA at final concentrations of 0.2, 2, 20 and 200 ng/ml, as well as sera from mice infected with S. japonicum on day 3, 7, 21, 28, 35 and 42 post-infection. Results The optimized RPA reaction system utilized final concentrations of 50 ng/μl for T4 recombinase Y, 440 ng/μl for T4 recombinase X, 200 ng/μl for T4 single-stranded binding protein and 120 ng/μl for creatine kinase, and the optimized reaction condition was 40 ℃ for 30 min. Sensitivity evaluation of established RPA-CRISPR/Cas12a method indicated the reaction products were positive when the S. japonicum genomic DNA content exceeded 10^-5 ng. Specificity evaluation indicated the reaction product of S. japonicum genomic DNA was positive, and the reaction products of S. mansoni, P. westermani and C. sinensis genomic DNA were negative. The results of mice dummy positive sera showed that the reaction products of S. japonicum genomic DNA concentrations of 2, 20 and 200 ng/μl were positive, and the reaction product of 0.2 ng/ml was weakly positive. The reaction products of sera from mice infected with S. japonicum on day 28 and 42 post-infection were positive, and the reaction product of sera on day 35 was weakly positive. Conclusion The study successfully established a one-tube RPA-CRISPR/Cas12a method for detecting the SjCHGCS20 sequence. The method has a lowest detectable limit of 10^-5 ng for S. japonicum genomic DNA, with no cross-reaction with genomic DNA from S. mansoni, P. westermani or C. sinensis. It is rapid, sensitive, specific and simple to operate, allowing for visual observation of results under blue light, and holds potential for field diagnosis of schistosomiasis japonica.

Analysis on Taenia infection and serum cysticercus antibody in population and their risk factors in Dali, Yunnan Province

WANG Wenya, LIU Jianfeng, ZHANG Mizhen, QIAN Peijun, LIU Liu, ZHAO Luyuan, LI Kerong, ZHANG Dongqi, LIU Yuhua, LIU Hongkun, CHEN Shaorong, ZHOU Changhai, QIAN Menbao, LI Shizhu

2024, 42(5):  615-622.  doi:10.12140/j.issn.1000-7423.2024.05.008

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Objective To analyze the epidemic characteristics and influencing factors of Taenia infection and cysticercus antibody in the residents of Dali, Yunnan, to provide a scientific basis for prevention and control of taeniasis and cysticercosis. Methods From October to November 2023, a survey was conducted in Dabaguan Village, Huoshan Village, Jingang Village and Qingshuigou Village in the historical endemic area of taeniasis and cysticercosis in Dali Prefecture. Permanent residents aged over 3 years were enrolled in the survey using the method of cluster random sampling. A questionnaire survey was conducted to collect information including whether there was a history of proglottid excreted, subcutaneous nodules, symptoms of epilepsy and severe headache, as well as the dietary habits, hygiene behaviors and living environment. Fecal samples were collected, and examined by the modified Kato-Katz method (two slide-reading for each sample). For residents with positive fecal test results or with a history of excreting proglottids in the past year, pharmacological diagnostic deworming was conducted using the areca-pumpkin seed method. Those positives in fecal examination and found expelled worms were considered as the infected persons and used to calculate the infection rate. Serum cysticercus specific antibodies were detected using cysticercus (Taenia solium) IgG antibody detection kit. The chi-square test, Fisher’s exact probability method and logistic regression analysis were used to analyze the influencing factors of Taenia infection and serum cysticercus antibody positivity in the population. Results A total of 1 842 people from the 4 villages participated in the questionnaire survey, and 39 people (2.1%) had excreted proglottids in the past year. Fecal examination was performed on 1 533 people, and 25 people (1.6%) were found positive. A total of 39 people received diagnostic deworming, and 33 people expelled complete worms (1 person positive in fecal examination did not expel the parasite), and the infection rate was 1.8% (34/1 842). The infection rates in the Jingang, Huoshan, Dabaguan and Qingshuigou villages were 2.1% (10/475), 1.3% (6/450), 2.7% (12/450) and 1.3% (6/467), respectively, with no significant difference (χ² = 3.31, P > 0.05). The infection rate was 2.6% (22/848) in males and 1.2% (12/994) in females (χ² = 4.90, P < 0.05). The infection rates of the 30-59 age group (2.8%, 24/862), Miao (1/2), and farmers and herdsmen (3.0%, 28/944) were the highest in each group (χ² = 10.10, P < 0.01; Fisher = 26.04, P < 0.01; χ² = 13.47, P < 0.01). The positive rate of serum cysticercus antibody was 9.2% (82/887). Positive rates of serum antibodies in Qingshuigou, Dabaguan, Huoshan and Jingang villages were 12.7% (20/157), 4.7% (11/236), 10.4% (26/251) and 10.3% (25/243), respectively, with significant differences (χ² = 8.88, P < 0.05). The positive rates in males were 9.2% (36/393) and 9.3% (46/494) in females (χ² = 0.01, P > 0.05). The serum antibody positive rates in the groups aged 60 years and above (2.8%, 24/862), farmers and herdsmen (11.2%, 61/546) and illiterate people (15.0%, 35/234) were the highest in each group (χ² = 8.66, 12.37, 6.47, P < 0.05 or 0.01). The results of univariate analysis showed that gender, age, nationality, occupation and consuming raw pork products (raw pig skin, pork and liver) were the influencing factors of Taenia infection (χ² = 4.86, 10.10, 26.04, 13.48, 5.72, all P < 0.05). There was a positive correlation between the frequency of consuming raw pork products per month and Taenia infection rate[OR = 1.135, 95% CI: (1.023, 1.259)]. The age, education level, occupation, use of tap water and consuming raw pork products were influencing factors for the positive rate of serum cysticercus antibody (χ² = 8.66, 12.37, 6.47, 3.58, 2.20, all P < 0.05). The results of multivariate analysis showed that males [OR = 2.047, 95% CI: (1.001, 4.189) ] and eating raw pork products [OR = 4.986, 95% CI: (1.166, 21.321) ] were the risk factors for Taenia infection. The low education level [OR = 2.051, 95% CI: (1.183, 3.557)] was a risk factor for positive serum cysticercus antibody, while using tap water [OR = 0.320, 95% CI: (0.124, 0.824) ] was a protective factor for positive serum cysticercus antibody. Conclusion Considerably high Taenia infection rate and positive rate of serum cysticercus antibody are observed in the residents of Dali, Yunnan. Eating raw pork products and male gender are the risk factors for Taenia infection; using tap water and high education level are the protective factors for positive serum cysticercus antibody.

Investigation of intestinal parasitic infections among the population in areas inhabited by Bulang nationality in Yunnan Province

CHEN Meng, TAO Hong, LI Yanzhong, ZHANG Juan, ZHOU Xiaomei

2024, 42(5):  623-628.  doi:10.12140/j.issn.1000-7423.2024.05.009

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Objective To investigate the current status of intestinal parasitic infections in areas inhabited by Bulang nationality in Yunnan Province, and to provide a scientific basis for formulating prevention and control strategies. Methods In 2015 and 2023, the survey was carried out in Bulang Township, Menghai County, Xishuangbanna Dai Autonomous Prefecture, Yunnan Province using cluster random sampling, and the survey subjects were permanent residents of all ages who lived for more than 6 months. The fecal samples and information of survey subjects such as gender, age and education level were collected. The modified Kato-Katz thick smear method (two slides per sample) was used to detect and count the eggs of intestinal worms, and each fecal sample was cultured to detect the hookworm used tube-filter paper culture method. Intestinal protozoa were detected by iodine staining smear method. Eggs, cysts or hookworms detected during fecal examination were judged to be intestinal parasitic infections, and the intensity of soil-transmitted nematode was determined. The population data of the two surveys were combined as the standard population to standardize the infection rate. The chi-square test was used for the comparison of the standardized infection rate. Results In 2015, a total of 800 people were surveyed in areas inhabited by Bulang nationality in Yunnan Province, and 564 people were positive for parasitic infection, with a standardized infection rate of 71.09%. In 2023, a total of 1 128 people were surveyed, and 203 people were positive for parasitic infection, with a standardized infection rate of 17.84%. The standardized infection rate of parasites in 2023 was lower than that in 2015 (χ² = 1 107.636, P < 0.01). The standardized infection rates of intestinal worms in 2023 was 12.17%, which was lower than that in 2015 (69.36%) (χ² = 1 304.222, P < 0.01). The standardized infection rates of Ascaris lumbricoides, hookworm and Trichuris trichura in 2023 (2.78%, 6.76% and 3.13%) were lower than those in 2015 (54.67%, 25.86% and 30.22%) (χ² = 1 266.428, 258.666, 510.527, all P < 0.01). In 2015 and 2023, the intensity of A. lumbricoides, hookworm and T. trichura were mainly mild infected. In 2015 and 2023, the standardized infection rates of intestinal protozoa were 5.21% and 6.39%, respectively, with no significant difference (χ² = 2.518, P > 0.05). The standardized infection rate of Blastocystis hominis in 2023 (3.11%) was higher than that in 2015 (1.04%) (χ² = 20.424, P < 0.01). The standardized infection rates of females and males were 75.47% and 66.18% in 2015 and 20.53% and 14.92% in 2023, respectively. The standardized infection rates of females in 2015 and 2023 were higher than those in males (χ² = 20.035, 10.416, both P < 0.01). The 20-29 year age group had the highest standardized infection rate of 23.53% in 2023, and there were statistically significant differences between the age groups (χ² = 14.683, P < 0.05); but no significant difference in 2015 (χ² = 13.211, P > 0.05). There was a significant difference in the standardized infection rate among the different education levels in 2015 (χ² = 21.108, P < 0.01), but no significant difference in 2023 (χ² = 2.007, P > 0.05). The standardized infection rate of Bulang nationality was 71.03%, which was higher than that of other nationalities (1.39%) (χ² = 155.480, P < 0.01) in 2015, and there was no significant difference between the standardized infection rate of Bulang nationality (17.73%) and other nationalities (20.83%) in 2023 (χ² = 0.456, P > 0.05). Conclusion In 2023, the infection rate of intestinal parasites in areas inhabited by Bulang nationality in Yunnan Province decreased significantly compared with 2015. The infection rate of intestinal worms decreased significantly, and the infection rate of intestinal protozoa increased.

Analysis on the surveillance of soil-transmitted nematode infection in Shanxi Province from 2019 to 2023

WANG Jingying, TIE Ping, ZHENG Yuhua, BAI Yongfei, WANG Ting, WANG Santao

2024, 42(5):  629-634.  doi:10.12140/j.issn.1000-7423.2024.05.010

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Objective To analyze the surveillance of soil-transmitted nematode (STN) infection in Shanxi Province from 2019 to 2023, to understand its current status and trends, providing scientific basis for developing control strategies. Methods From 2019 to 2023, in accordance with the requirements of the National Surveillance Program and Implementation Guideline for Clonorchiasis and Soil-transmitted Nematodes, in different regions of Shanxi Province, 14, 18, 15, 18, and 18 counties were selected for the surveillance, respectively. Each selected surveillance county was divided into five geographic sectors (east, south, west, north, and center). From each sector, one township (town, street) was randomly selected, and subsequently, one administrative village was chosen from each selected township for the surveillance. In each selected village, the participants were cluster sampled, with no less 200 villigers for the survey. Fecal samples of no less than 30 g were collected from the participants, and examined microscopically using the modified Kato-Katz thick smear method (two slide-reading, each sample) for STN eggs to calculate the infection rate. Hookworm egg positive fecal samples were further examined using the test tube-filter paper cultivation method to identify the parasite species. Additionally, microscopic-detection of Enterobius vermicularis eggs was conducted in children aged 3 to 9 using the transparent adhesive paper anal swab method. χ² test was employed to compare infection rates. Results From 2019 to 2023, the infection rates of STN in the surveillance population in Shanxi Province were 0.05% (7/14 189), 0.05% (9/18 019), 0.12% (18/15 024), 0 (0/18 109), and 0.11% (21/18 383), respectively. There were statistically significant differences in infection rates between years (χ² = 26.464, P < 0.01). During this period, the infection rates of Ascaris lumbricoides were 0.02% (3/14 189), 0.01% (1/18 019), 0 (0/15 024), 0 (0/18 109), 0.04% (8/18 383); and the infection rates of E. vermicularis were 0.02% (3/14 189), 0.04% (8/18 019), 0.12% (18/15 024), 0 (0/18 109), 0.07% (13/18 383), respectively. The infection rates of both A. lumbricoides and E. vermicularis showed statistically significant differences between years (χ² = 13.586 and 27.674, both P < 0.01). The Trichuris trichiura infection rate was 0.007% (1/14 189) in 2019, and 0 in the subsequent years. Notably, no hookworm infections were found from 2019 to 2023. Regarding gender, the infection rates for STNs in male and female were 0.07% (30/40 707), and 0.06% (25/43 017), respectively, with no statistically significant difference (χ² = 0.773, P > 0.05). In age groups, the highest infection rate of STNs was 0.30% (30/9 913) in the 3-9 year-old group, followed by 0.05% (8/14 828) in the 50-59 year-old group, and 0.05% (3/5 938) in the 20-29 year-old group. There were statistically significant differences in the infection rates between different age groups (χ² = 98.579, P < 0.01). Among different occupations, scattered children had the highest average infection rate (0.90%, 6/665), followed by nursery children (0.36%, 17/4 707). The differences in infection rates among different occupations were statistically significant (χ² = 64.542, P < 0.01). In terms of literacy levels, preschool children had the highest average infection rate of 0.42% (23/5 433), followed by elementary school at 0.07% (16/24 103). There was a statistically significant difference in infection rates among different literacy levels (χ² = 57.218, P < 0.01). In 2019—2023, Shanxi Province completed monitoring in 79 different surveillance counties in 11 cities, with positive infections found in 17 of these counties. Xia County of Yuncheng City had the highest infection rate of 1.00% (10/1 000), followed by Wenshui County of Lvliang City, and Lingshi County of Jinzhong City, both at 0.60% (6/1 003) 0.60% (6/1 003). The differences in infection rates among the different surveillance counties were statistically significant (χ² = 29.741, P < 0.05). Conclusion The infection rate of STN in Shanxi Province from 2019 to 2023 showed a trend of low-level prevalence, with E. vermicularis being the primary parasite. It is necessary to further strengthen the comprehensive control of STN infections, particularly among key populations such as children and the elderly.

Epidemiological characteristics of imported malaria in Fujian Province from 2014 to 2023

XIAO Lizhen, OUYANG Rong, CHEN Zhuyun, LIN Yaoying, ZHENG Dan, XIE Hanguo

2024, 42(5):  635-641.  doi:10.12140/j.issn.1000-7423.2024.05.011

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Objective To analyze the epidemiological characteristics of imported malaria in Fujian Province from 2014 to 2023, so as to provide scientific evidence for consolidating the achievements of malaria elimination. Methods The information of reported malaria cases in Fujian Province from January 1, 2014 to December 31, 2023 were collected from the data system of National Information System for Infectious Disease Surveillance and the National Information System for Parasitic Disease Control and Prevention. Of the collected information, malaria species, infection sources, three-dimentional distribution, medical consultation and diagnosis were analyzed statistically. Chi-square test or ANOVA were used for comparison of differences. Results A total of 825 imported malaria cases were reported in Fujian Province during 2014-2023, all of them were imported cases from abroad. The laboratory diagnosis found that 607 cases were infected with Plasmodium falciparum, 129 P. vivax, 62 P. ovale, 23 P. malariae and 4 mixed infection of P. falciparum and P. ovale. From 2014 to 2019 and 2023 (before or after the COVID-19 epidemic) and from 2020 to 2023 (during the COVID-19 epidemic), the proportion of falciparum malaria cases was 74.7% (567/759) before and after the COVID-19 epidemic, and 60.6% (40/66) during the COVID-19 epidemic. The proportion of other cases (vivax malaria, ovale malaria, quartan malariae and mixed infection) was 25.3% (192/759) before and after the COVID-19 epidemic and 39.4% (26/66) during the COVID-19 epidemic. The difference of diagnosis classification proportion of reported cases was statistically significant between the COVID-19 epidemic period and before or after the COVID-19 epidemic (χ² = 6.207, P < 0.05). The reported cases were imported from 42 countries in four continents. The malaria cases originated from 32 countries in Africa (748 cases), 7 countries in Asia (51 cases), 1 country in Oceania (24 cases) and 2 countries in South America (2 cases). Malaria cases were reported from all 9 cities, mainly from Fuzhou (75.2%, 620/825). A total of 56.0% (47/84) of the counties (cities and districts) reported imported malaria cases, with Gulou District of Fuzhou accounting for 71.1% (587/825). There were cases reported in every month, with an average of 6.88 cases per month. The monthly average number of reported cases during the COVID-19 epidemic was 1.83, which was lower than that before or after the COVID-19 epidemic (9.04 cases) (t = 12.229, P < 0.05). Among the reported cases, males accounted for 90.1% (743/825), females for 9.9% (82/825), with a male-to-female ratio being 9.1 : 1. The cases were mainly at the age of 30-39 years and 40-49 years (60.4%，498/825). Cases were reported from 89 institutions. The median interval between onset and initial visit was 2 days. Most of the malaria cases (74.3%, 613/825) visited doctor 3 days after outset of disease, but 4.9% (40/825) of the cases consulted doctor over 7 days after outset. The proportion of patients seeking treatment within 3 days of onset of falciparum malaria (77.3%, 469/607) was higher than that of other classified cases (66.0%, 144/218) (χ² = 10.547, P < 0.05). The median time interval between initial diagnosis and diagnosis was 0 days. 83.4% (688/825) of the cases were diagnosed as malaria within 3 days; 5.5% (45/825) of the cases spent more than 7 days of time span from initial visit to confirmed diagnosis. The proportion of falciparum malaria cases diagnosed within 3 days (86.3%, 524/607) was higher than that of other classified cases (75.2%, 164/218) (χ² = 14.262, P < 0.05). A total of 43 severe malaria cases were reported in Fujian Province from 2014 to 2023, accounting for 5.2% (43/825) of the total reported malaria cases. Among them, 1 case was a mixed infection of P. falciparum and P. ovale, and the other 42 cases were infected with P. falciparum. Conclusion All malaria cases reported in Fujian Province from 2014 to 2023 were imported cases. Imported malaria is the key target in prevention and control work in Fujian Province in the future. It is inperative to further strengthen health education for inbound and outbound persons, and the technical capability of medical personnel in diagnosis.

REVIEWS

Research progress on the interaction between intestinal nematodes and intestinal flora

ZHANG Zhilan, YIN Sixuan, LV Fangli

2024, 42(5):  642-647.  doi:10.12140/j.issn.1000-7423.2024.05.012

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The intestinal flora is an important component of human intestinal microenvironment, playing important roles in digestion and absorption, nutrient metabolism, defense against pathogenic infections and regulation of autoimmune diseases. There are complex interactions between parasites residing in the host’s gut and the host’s gut microbiota, influencing the health of the host’s intestine and the onset and progression of related diseases. This article reviews the research progress on the interaction between host intestinal nematodes and intestinal flora, to provide scientific basis for a deeper understanding of the pathogenic mechanisms of related diseases.

Research advances on the effects of helminth infection on bile acid metabolism

ZHANG Ying, WANG Yugui, SHI Zhiqi, WANG Shuai

2024, 42(5):  648-652.  doi:10.12140/j.issn.1000-7423.2024.05.013

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Parasitic helminths are widely distributed globally and have a significant impact on animal husbandry and public health. Helminth infection can affect metabolic and immune processes in the host. The latest research shows that helminth infection significantly alters host bile acid metabolism, which in turn directly or indirectly affects the host’s health and immune homeostasis. This article reviews the advances in research on the effects of nematodes, tapeworms, and flukes on host bile acid metabolism to provide new research ideas for the diagnosis, treatment, and prevention of related diseases.

Research advances on host cell-autonomous immunity against Toxoplasma gondii

LI Shiyu, LI Jing, LU Shaohong, ZHENG Bin

2024, 42(5):  653-658.  doi:10.12140/j.issn.1000-7423.2024.05.014

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Toxoplasma gondii is one of the most common infectious agents in humans and animals, causing zoonotic toxoplasmosis, which poses a serious hazard to human health and animal husbandry production. Cell-autonomous immunity is an intrinsic immune response that occurs within individual cells to limit the replication and spread of pathogens (e.g., bacteria, viruses, parasites, etc.) that invade the cells. After infection of host cells by T. gondii, cell-autonomous immunity is mainly regulated by interferon-induced indoleamine 2,3-dioxygenase (IDO), inducible nitric oxide synthase (iNOS), GTPase family enzymes, autophagy-related proteins, and ubiquitylation products, all of which play an important role in host cell resistance to T. gondii. In this paper, we reviewed the research advances on the autonomous immune response of host cell against T. gondii, aiming to provide new directions for the prevention and treatment of toxoplasmosis.

SHORT COMMUNICATIONS

Analysis of the current status of Clonorchis sinensis infection in Xinfeng County, Jiangxi Province from 2020 to 2022

LAN Weiming, ZENG Yufeng, QIU Tingting, ZHOU Changhai, CHEN Zhe, GE Jun, ZENG Xiaojun, ZHU Tingjun

2024, 42(5):  659-663.  doi:10.12140/j.issn.1000-7423.2024.05.015

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In order to understand the current situation of Clonorchis sinensis infection in Xinfeng County, Jiangxi Province, Xinfeng County was divided into 5 sectors (east, west, south, north and central) according to the geographical location. From each sector, 1 administrative village in 1 township (town) was selected as surveillance site, and no less than 200 permanent residents aged 3 or above were selected by cluster random sampling from each surveillance site in every year. Fecal samples were collected and examined for eggs of C. sinensis by modified Kato-Katz thick smear method (two slides per sample). The infected cases were treated with praziquantel (75 mg/kg) for 2 days, and examined for eggs of C. sinensis one month after treatment. The infection rate was compared by Chi-square test. The total infection rate of C. sinensis was 6.43% (202/3 142) in Xinfeng County from 2020 to 2022, and the annual infection rates were 6.09% (61/1 002), 5.11% (52/1 017) and 7.93% (89/1 123), respectively (χ² = 7.30, P < 0.05). The infection rate of males (8.30%, 134/1 614) was higher than females (4.45%, 68/1 528) (χ² = 19.36, P < 0.01). The infection rates of the elderly groups over 40 years old (7.87%, 168/2 134) was higher than that of the younger age groups (3.37%, 34/1 008) (χ² = 23.04, P < 0.01). The groups with the highest infection rates were college and above in 2020 (1/8), junior high school in 2021 (6.02%, 15/249) and illiteracy in 2022 (11.36%, 30/264), respectively. The infected cases originated from 176 households, and the proportions of households with 1, 2 and 4 infected cases were 87.50% (154/176), 11.36% (20/176) and 1.14% (2/176), respectively. After praziquantel treatment, 197 infected cases achieved negative in fecal test, among them 2 cases turned positive after 1 year and 10 cases turned positive after 2 years. The reinfection rate was 6.09% (12/197). The results indicated that the prevention and control situation for clonorchiasis in Xinfeng County, Jiangxi Province was still severe. The infection rates were relatively high in some administrative villages, with familial clustering of infection and individual reinfection.

Analysis of polymorphism of Pfhrp2 gene in imported Plasmodium falciparum in Henan Province in 2020

LI Suhua, JI Penghui, WANG Dan, WANG Hao, ZHOU Ruimin

2024, 42(5):  664-667.  doi:10.12140/j.issn.1000-7423.2024.05.016

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To analyze the deletion and polymorphism of exon 2 region of Plasmodium falciparum histidine rich protein 2 (Pfhrp2) gene from the blood samples of the imported falciparum malaria cases in Henan Province in 2020. The information and blood samples of the imported falciparum malaria cases confirmed by Henan Provincial Malaria Diagnosis Reference Laboratory in 2020 were collected. Nested PCR was performed to amplify Pfhrp2 exon 2 used DNA extracted from blood samples, and the positive amplification products were sequenced. A total of 37 blood samples of confirmed imported falciparum malaria cases were collected. 35 sequences of Pfhrp2 exon 2 were amplified successfully by nested PCR and the positive rates was 94.59% (35/37). 28 of 35 positive amplification products were sequenced successfully, with an average length of 822 bp. Twelve kinds of repeat types were found in the amino acid chains encoded by Pfhrp2 exon 2. All the amino acid chains started with types 1 (AHHAHHVAD) and ended with types 12 (AHHAA), and contained type 1, 2, 4, 11 and 12. The proportion of other repeat types in descending order were type 13 (96.43%, 27/28), 3 (92.86%, 26/28), 7 (82.14%, 23/28), 8 (64.29%, 18/28), 6 (60.71%, 17/28), 5 (46.43%, 13/28) and 9 (32.14%, 9/28), respectively. The results of this study showed that there was a high diversity of Pfhrp2 exon 2 in imported Plasmodium falciparum from Henan Province in 2020, which might cause different results of rapid diagnostic tests.

Phylogenetic and sequence analysis of mitochondrial nad1 of Metorchis orientalis strains from different geographic regions

CHEN Yunhong, XIE Xianliang, LI Yanrong, CHEN Baojian, LIN Yaoying, YU Wenwu, YU Haifeng

2024, 42(5):  668-671.  doi:10.12140/j.issn.1000-7423.2024.05.017

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To examine the genetic variation and phylogenetic relationship of nicotinamide adenine dinucleotide dehydrogenase subunits 1 (pnad1) of Metorchis orientalis from different geographic regions. The quail were infected with M. orientalis metacercaria isolated from Pseudorasbora parvain in Fujian Province and Jiangxi Province, respectively. Adult worms were collected after 30 days, and the mitochondrial genomic DNA was extracted from 3 adults in each region. The pnad1 was amplified from M. orientalis by PCR and sequenced. The obtained sequences were compared with the mitochondrial pnad1 sequences from Heilongjiang (GenBank: KY232061, KY232066, KY232071) and published sequences of the same genus using BioEdit software and the species-specific and inter-species differences were analyzed. The phylogenetic distance of different regions was calculated by MEGA 11.0 software, and the phylogenetic tree of opisthorchidae based on mitochondrial pnad1 sequence was constructed by the adjacency method. The results showed that 500-bp bands were amplified in the mitochondria of the M. orientalis from Fujian and Jiangxi, and the mitochondrial pnad1 gene was 523 bp. The results of sequence comparison showed that the intraspecific genetic variation rate of pnad1 sequence was 0-0.8%, and the interspecific genetic variation rate was 12.6%-13.9%. The constracted phylogenetic tree based on the pnad1 gene sequences showed that M. orientalis isolates from Fujian, Jiangxi and Heilongjiang were clustered together, M. xanthosomus (GenBank: KT740991) and M. bilis (GenBank: NC079698) were clustered in a branch. It was concluded that the mitochondrial pnad1 sequence is relatively conserved within species and varies greatly between species, which can be used as a marker for studying interspecific genetic variation.

Surveillance and analysis of human intestinal helminth infection in Shandong Province in 2023

LV Wenxiang, BU Cancan, LI Yuejin, WANG Longjiang, WANG Yongbin, KONG Xiangli, ZHANG Benguang, YAN Ge, XU Yan

2024, 42(5):  672-675.  doi:10.12140/j.issn.1000-7423.2024.05.018

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To analyze the human intestinal helminth infection in Shandong Province and provide a scientific basis for developing parasite control strategies and measures. The survey was conducted according to the requirements of the National Surveillance Program for Key Human Parasitic Diseases, and 14 survey sites were selected as the investigation points in 9 cities, including Jinan, Qingdao, Weifang, Jining, Linyi, Dezhou, Liaocheng, Binzhou and Heze. Each survey point was assigned to 5 regions in the east, south, west, north and middle according to geographical orientation, and 1 administrative village from 1 neighbourhood was selected in each region for monitoring. At least 200 permanent residents over the age of 3 years were selected from each village. The modified Kato-Katz thick smear method was used to examine parasite eggs (two slide-reading for each sample). Children aged 3 to 9 years in each village were also tested with the cellophane tape anal swab method to examine Enterobius vermicularis eggs. SPSS 21.0 software was used for statistical analysis, and the difference between infection rates was compared with χ² test. A total of 14 507 people were investigated, 45 people were found to be infected with intestinal helminth, the total infection rate was 0.31% (45/14 507). The infection rates of Ascaris lumbricoides, Trichuris trichura, Clonorchis sinensis and E. vermicularis were 0.01% (2/14 507), 0.30% (43/14 507), 0.01% (2/14 507), 0.82% (15/1 832), respectively. The infection rate of intestinal helminth was the highest in Linyi City (4.16%, 42/1 010). The infection rate of Jining City was 0.10% (1/1 014). The infection rates in Qingdao and Weifang were 0.05% (1/2 050, 1/2 115). The difference was statistically significant (χ² = 196.906, P < 0.01). The rates of E. vermicularis infection in Dezhou, Liaocheng, Binzhou and Heze were 0.96% (2/208), 3.00% (3/100), 1.57% (6/382), 2.78% (4/144) respectively, with no significant difference (χ² = 2.536, P > 0.05). The infection rate of intestinal helminth was 0.28% (19/6 805) in males and 0.36% (28/7 702) in females. The infection rate of intestinal helminth in the 19-40 years old group was the highest at 0.40% (12/2 988). The infection rates of intestinal helminth were the highest in primary school and middle school education levels, both of which were 0.41% (18/4 427, 16/3 933). There were no statistically significant differences in intestinal helminth infection rate between different genders, different age groups and different educational levels (χ² = 0.796, 2.837, 5.251; P > 0.05). The highest intestinal helminth infection rate was 0.56% (11/1 970) in students, and there was a significant difference in intestinal helminth infection rate among different occupations (χ² = 23.847, P < 0.01). The infection rate of human intestinal helminth in Shandong Province is at a low level, but there is still a risk of transmission in some areas, and surveillance and prevention should be continued.

Clinical analysis of two cases of imported visceral leishmaniasis in infants in Yunnan Province

SHI Shulan, LI Cuiying, ZHANG Qian, LI Rongjie, KUI Liyue, FAN Mao

2024, 42(5):  676-678.  doi:10.12140/j.issn.1000-7423.2024.05.019

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This retrospective analysis reviews the laboratory and clinical data of two infants with visceral leishmaniasis treated at Kunming Children’s Hospital from January 2021 to December 2022. Both patients were female from Yunnan, and 9 months and 2 years old, respectively, with a history of residence in endemic areas (Gannan Tibetan Autonomous Prefecture, Gansu Province, and Kuqa City, Xinjiang Uygur Autonomous Region) and a history of sandfly bites. Symptoms included irregular fever (39.7 ℃ to 40 ℃), rash, and pallor. Blood routine tests showed decreased white blood cells (3.92 × 10⁹/L, 3.34 × 10⁹/L), hemoglobin (98 g/L, 79 g/L), and platelets (49 × 10⁹/L, 57 × 10⁹/L), with elevated C-reactive protein (200 mg/L, 58.19 mg/L), procalcitonin (2.29 ng/ml, 1.38 ng/ml), and ferritin (69 445 µg/L, 2 800 µg/L). rK39 antibody tests were positive. Bone marrow smears revealed Leishmania amastigotes. The second case,through comprehensive pathogen metagenomic next-generation sequencing, identified 32 026 sequences of Leishmania donovani. Abdominal ultrasound showed hepatosplenomegaly in both patients. After diagnosis, sodium stibogluconate was administered at 240 mg/kg divided into 6 doses, with one intravenous injection per day. It resulted in a cure, with no abnormalities at six months and one year follow-up. Visceral leishmaniasis in children can have atypical symptoms. If there is prolonged irregular fever, clinicians should inquire in detail about the epidemiological history of visceral leishmaniasis. Metagenomic sequencing may be necessary to avoid misdiagnosis or missed diagnosis if no pathogens are found in the bone marrow.

Investigation of common Culicoides biting midges in human settlements in Fujian Province

CHEN Zhuyun, XIAO Lizhen, OUYANG Rong, XIE Hanguo

2024, 42(5):  679-683.  doi:10.12140/j.issn.1000-7423.2024.05.020

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To investigate the population distribution of common Culicoides biting midges in human settlements in Fujian Province, 19 surveillance sites were set up at different environment in the human settlements (human house, field, livestock houses and mangroves near houses) of 9 cities in Fujian Province from June, 2022 to September, 2023, and the midges were collected by light trap. The species of midges were identified by morphological classification and molecular identification of cylochrome C oxidase subunit 1 (cox1) gene. The database was established by Microsoft Excel 2007 software and SPSS 26.0 software was used to perform the statistical analysis. The degree of linear correlation between meteorological conditions and the number of Culicoides was analyzed by Pearson correlation analysis. A total of 7 862 Culicoides were collected, which were identified as 13 species belonging to 6 subgenera. Twelve species were identified by morphological classification, and 1 species was identified as C. circumbasalis by molecular identification. C. arakawae had the broadest distribution, being caught in all 19 surveillance sites. The number of Culicoides caught in human house, field, livestock houses and mangroves accounted for 7.10% (558/7 862), 12.79% (1 006/7 862), 30.44% (2 393/7 862) and 49.67% (3 905/7 862) of the total, respectively. C. arakawae was the dominant species of human house (85.84%, 479/558), field (84.00%, 845/1 006) and livestock houses (87.67%, 2 098/2 393), and C. circumbasalis was the dominant species of mangroves (99.74%, 3 895/3 905). C. circumbasalis was consistently captured in the mangroves throughout the entire year, with a notable peak in July (52.32%, 2 038/3 895). The number of catched C. circumbasalis showed a positive correlation with the average monthly temperature (r = 0.628, P < 0.05), but no correlation with mean wind speed or rainfall (r = -0.316, -0.073, both P > 0.05). The results of this investigation showed that the population of Culicoides in human settlements was diverse and abundant in Fujian Province. C. arakawae had the broadest distribution, being the dominant species of human house, field and livestock houses, and C. circumbasalis was the dominant species of mangroves.

CASE REPORT

A case of secondary myelofibrosis in visceral leishmaniasis

YANG Mei, XIANG Baoyun, WEI Tao, CHI Yonge, LI Juan

2024, 42(5):  684-687.  doi:10.12140/j.issn.1000-7423.2024.05.021

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The case was a 15-year-old male patient from Linzhou, Henan province and he was a student. Due to “intermittent fever for more than half a month”, he went to a local municipal hospital on March 3, 2021. The blood routine test showed leukocyte was 2.60 × 10⁹/L, platelet was 65 × 10⁹/L and hemoglobin was 113 g/L. He was diagnosed with infectious fever with leukothrombopenia and splenomegaly. After anti-infection treatment, the symptoms were improved and he was discharged home. But the condition sometimes recurred and he was admitted to the hematology department of a provincial hospital because of “repeated fever with fatigue and pancytopenia for more than 1 year” on April 19, 2022. The blood routine test showed white blood cell was 1.79 × 10⁹/L, hemoglobin was 73 g/L and platelet was 54 × 10⁹/L. Ultrasound showed enlarged lymphnodes in the neck and axilla, large liver and spleen with widened splenic veins. Bone marrow smear showed hyperplasia was active, granulocyte series reduced, erythrocyte series was significantly active while mature erythrocytes clustered and arranged in row, and no megakaryocyte was found in this smear. Bone marrow biopsy showed extensive fibrosis, increasing proportion of erythrocyte series, many megakaryocyte cells and also showed T cells were easy to be seen with scattered individual large cells. Positron emission tomography-computed tomography (PET-CT) showed diffuse active in splenic metabolism and no abnormalities in hepatic metabolism. Then the patient was suspected to have a splenic lymphoma and was transferred to surgery for splenectomy + abdominal lymph node dissection. Pathology of the spleen showed moderate histiocytic hyperplasia and pathogen in the cytoplasm of the histocyte which was verified Leishmania donovani by gene sequencing. Pathology of the spleen also showed extramedullary hematopoietic. The patient was diagnosed with visceral leishmaniasis. The patient had a history of contact with dogs and the county where he lived had visceral leishmaniasis case report. He was admitted to the Fifth People’s Hospital of Anyang in June 27, 2022 and was began to be treated with sodium stibogluconate injection from that time. The admission examination showed emaciation, malnutrition and hepatomegaly that liver could be palpated 10cm below costal margin on right midclavicular line, moderate quality, no tenderness. Blood routine test showed that the white blood cell was 11.17 × 10⁹/L， hemogloubin was 10⁴ g/L and platelet was 245 × 10⁹/L. The result of rK39 kala-azar detect rapid test was positive. Abdominal CT showed a significantly enlarged liver. After treated with sodium stibogluconate injection for 1 course (0.6 g/d, 9 d), The temperature returned to normal and the general condition improved, but Leishmania amastigote was visible in his bone marrow smear. So another course sodium stibogluconate injection (0.6 g/d, 9 d) was given to him. After the second course’s treatment, the bone marrow smear was reexamined and no Leishmania amastigote could be found. The paitent was recovered and discharged home. He felt well and the abdominal CT showed liver was somewhat reduced and bone marrow trephine biopsy performed had returned to normal and bone marrow smear was not found Leishmania amastigote about 10 months after initiation of the therapy.