Table of Content

28 February 2020, Volume 38 Issue 1

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EXPERT VIEWPOINT

Suggestions for standardizing the process of malaria diagnosis and treatment during the epidemic control of coronavirus disease(COVID-19)

ZOU Yang, ZHENG Yi-shan, CAO Jun

2020, 38(1):  1-4.  doi:10.12140/j.issn.1000-7423.2020.01.001

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At the end of 2019, the epidemic of coronavirus disease 2019 (COVID-19) brought new challenges to the diagnosis and treatment of other diseases such as malaria. If malaria patients are misdiagnosed, the illness may become severe or even life-threatening. Furthermore, if improperly protected and handled against the epidemic, not only malaria patients may be infected with the novel coronavirus during the process seeking for medical care, but medical staff will also face potential high risks. Therefore, to provide reference for health care workers, this paper focuses on how to perform good practice for malaria diagnosis and treatment under the current COVID-19 epidemic, and provides recommendations for the standardized diagnosis and treatment process of malaria patients.

ORIGINAL ARTICLES

National survey on the current status of important human parasitic diseases in China in 2015

CHEN Ying-dan, ZHOU Chang-hai, ZHU Hui-hui, HUANG Ji-lei, DUAN Lei, ZHU Tin-jun, QIAN Men-bao, LI Shi-zhu, CHEN Hong-gen, CAI Li, HU Tao, LEI Zheng-long, LI Hua-zhong, LI Zhong-jie, WANG Tian-ping, SUN Jun-ling, XU Bian-li, YAN Jun, YANG Yi-chao, ZENG Xiao-jun, WANG Guo-fei, ZANG Wei, WANG Ju-jun, ZHOU Xiao-nong

2020, 38(1):  5-12.  doi:10.12140/j.issn.1000-7423.2020.01.002

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Objective To understand and analyze the current status and endemic trends of important human parasitic diseases, and provide scientific basis for the formulation of control programs in China. Methods The survey was carried out in 31 provinces (autonomous regions/municipalities) (hereinafter referred to as "P/A/Ms", excluding Hong Kong, Macao and Taiwan regions) of China from 2014 to 2016, using stratified cluster random sampling method. In rural areas, survey sites were selected based on the strata of ecological function in the nationwide eco-mapping and farmer’s income level (high, moderate and low annual net income per capita) in the county, by sampling 4-36 counties per province and 2-4 natural villages each sampled county. Totally, 1 890 survey sites were sampled, from each of them 250 permanent residents were surveyed by fecal examination for helminthes infection using modified Kato-Katz thick smear method (one sample, two slide-readings), applying saline smear and iodine solution smear method for protozoa infection, and by anal transparent tape test to examine Enterobius vermicularis infection for the children aged 3-6 years. In urban areas, however, only human Clonorchis sinensis infection was surveyed by fecal examination. For this purpose, the country was categorized into five endemic regions (I-V) based on the C. sinensis infection prevalence by region. A total of 517 survey sites were sampled, by selecting 2-37 counties from each province of all endemic regions, and taking urban resident community as the survey site. From each site, 250 permanent residents were surveyed for C. sinensis infection. Results A total of 617 441 people were surveyed in 31 P/A/Ms, comprising 484 210 from rural areas and 133 231 from urban areas, among them, 20 351 were found infected with important parasites (detection rate, 3.30%). A total of 34 species of parasites were found, including 23 helminth species and 11 protozoan species. The weighted infection rate of important parasites was 5.96%, accordingly, having estimated 38.59 million infected people. More specifically, the weighted infection rate of helminths was 5.10%, and that of intestinal protozoa was 0.99%, estimating 6.42 million people being infected. The weighted infection rate of soil-transmitted helminthes (STHs) was 4.49%, estimating 29.12 million people infected. The infection rate of Enterobius vermicularis was 3.43% in children aged 3-6 years, estimating 1.55 million infected children. The weighted infection rate of Taenia spp. was 0.06%, estimating 0.37 million infected. The weighted infection rate of C. sinensis in the country was 0.47%, estimating 5.98 million infected people. The weighted infection rate of C. sinensis in rural areas and urban areas was 0.23% and 0.71%, respectively, estimating 1.52 million and 4.46 million people infected. The important parasite infection displays an obvious regional distribution. The highly endemic areas of STHs infection were mainly distributed in Sichuan, Hainan, Guizhou, Yunnan, Chongqing, Guangxi, Guangdong and Jiangxi; the endemic areas of C. sinensis were restrictedly distributed in Guangdong, Guangxi, Heilongjiang and Jilin; the Taenia infection was mainly distributed in Tibet; and the highly endemic areas of E. vermicularis infection in children aged 3-6 years were mainly located in Hainan, Jiangxi, Guangdong, Guangxi, Guizhou and Chongqing. Over 50% of intestinal protozoa-infected people were distributed concentratedly in west China including Tibet, Guizhou and Guangxi. Conclusion The infection rate of important parasites, especially STHs, has been reduced significantly in China, showing a low level of prevalence or sporadically distributed in most of areas, and the scope of endemic regions has been markedly reduced as well. However, severe infection still exists in some provinces or local areas. The important parasite infection displays evident regional distribution pattern. Medium- and high-prevalence areas of STHs are mainly distributed in two broad regions in south and southwest China. The endemic areas of C. sinensis infection are mainly distributed in another two broad regions in southern and northeastern part of China. Taenia infection remains distributed in Tibet. The highly endemic areas of E. vermicularis infection in children aged 3-6 years are concentrated located in the southern and southwestern China. The intestinal protozoa infection is characterized with higher infection occurred in some limited areas, mainly distributed in western provinces. The number of people infected with important parasites in rural areas remains high, thus, prevention and control in this regard is still an arduous task.

Analysis of nad1 gene polymorphisms of Echinococcus granulosus isolates from humans in Ali region of Tibet

WEI Yu-huan, LIU Hua, LI Wu-jun, ZHAO Hai, HU Yuan, CAO Jian-ping

2020, 38(1):  17-22.  doi:10.12140/j.issn.1000-7423.2020.01.003

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Objective To understand the dominant genotypes and genetic variations of Echinococcus granulosus isolates from humans in Ali region, in order to provide support for tracing the origin of E. granulosus and formulating strategies for prevention and control of echinococcosis in Ali region. Methods Specimens resected from hydatidosis patients in 2017 in a hospital in Ali area were used for DNA extraction. Mitochondrial NADH dehydrogenase 1 (nad1) gene was amplified by PCR. The PCR products were sequenced and analyzed by BLAST, ClustalX 1.83 and MEGA 7.0 softwares for homology and phylogenetic analyses. The nad1 gene sequences of E. granulosus in China was downloaded and the haplotypes were analyzed by DnaSP6. The haplotype network map of nad1 gene of E. granulosus in China was drawn by the NetWork software. Results A total of 80 specimens of hydatidosis were collected, and 38 resulted in a specific band around 550 bp after PCR amplification. Sequencing analysis revealed that E. granulosus in 4 cases had the G6 genotype, which was 99.81% homogeneous with the sequence of Mongolian origin (MH300971.1). The remaining 34 cases revealed G1 genotype of E. granulosus, which had a C-to-G mutation at position 535 as compared to the standard sequence (AF297617.1) and was 100% homogeneous with that of the Algerian origin (MG672293.1). Analysis of the 38 samples with MEGA 7.0 software showed that the bases T, C, A and G constituted 44.3%-46.5%, 7.7%-9.1%, 19.8%-21.5% and 25.7%-26.0%, respectively. The reported E. granulosus nad1 gene in China had 9 haplotypes, with a haplotype diversity of 0.47 and nucleotide diversity of 0.19. The haplotype network map showed that H4 was the main haplotype for nad1 gene in China. Conclusion G1 and G6 are the genotypes of E. granulosus nad1 gene in Ali region. The data analysis indicates that H4 is the major haplotype for E. granulosus nad1 gene in China.

Genetic polymorphisms of mitochondrial cox1 gene of Echinococcus spp. from rodents and livestock in Xinyuan County, Xinjiang and Shiqu County, Sichuan

WANG Xu, HOU Yan-yan, WANG Ying, WANG Zheng-huan, XUE Chui-zhao, ZHANG Yin, LIU Bai-xue, HAN Shuai, ZHENG Can-jun, WU Wei-ping

2020, 38(1):  22-29.  doi:10.12140/j.issn.1000-7423.2020.01.004

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Objective To understand the genetic polymorphisms of cytochrome c oxidase subunit 1 (cox1) gene in mitochondria of Echinococcus spp. in Xinyuan County, Xinjiang and Shiqu County, Sichuan, and discuss Echinococcus transmission directions and dynamics in China. Methods From 2014 to 2018, rodents were captured in Xinyuan County (Xinjiang) and Shiqu County (Sichuan) to collect liver tissues, meanwhile lesioned tissue samples of livestock were collected. DNA was extracted from the livers and lesioned tissues, and underwent PCR to amplify part of Echinococcus mitochondrial cox1 gene. PCR products were sequenced, and the sequencing results were aligned in NCBI database to identify the Echinococcus species. The sequences were edited by Clustal X2 and MAGE 7 softwares, and diversity and neutrality indices were calculated by DnaSP v5 and Arlequin 3.5. Genetic diversities were compared between the two regions. By using E. shiquicus as an outgroup, cox1 sequence-based Bayesian phylogenetic trees were established with MrBayes 3.2.4 software. The haplotype networks were drawn for cox1 genes of Echinococcus spp. using Network 5.0 to analyze haplotype structure. Results One hundred and twenty-two Microtus arvalis were captured and 5 lesioned tissues of sheep were collected in Xinyuan County (Xinjiang); while in Shiqu County (Sichuan), 144 Lasiopodomys fuscus, 44 M. limnophilus and 135 Ochotona curzoniae were captured and 4 lesioned tissues of yaks were collected. A total of 40 sequences of E. granulosus cox1 genes (26 from Xinyuan, and 14 from Shiqu) were amplified from lesioned tissues of 5 sheeps and 4 yaks. Fifty-nine sequences of E. multilocularis mitochondrial cox1 genes (20 from Xinyuan, and 39 from Shiqu) were amplified in 8 M. arvalis, 14 L. fuscus, 5 M. limnophilus and 2 O. curzoniae. All PCR products had a length of 875 bp. E. granulosus showed a higher degree of genetic differentiation (Fst = 0.088 63) between the 2 regions than E. multilocularis (Fst = 0.000 88). The genetic diversity of E. granulosus was lower in Xinyuan (0.002 58 ± 0.000 46) than in Shiqu (0.005 88 ± 0.000 58), while E. multilocularis had higher genetic diversity in Xinyuan (0.002 28 ± 0.000 46) than in Shiqu (0.001 37 ± 0.000 30). The Bayesian phylogenetic tree showed that in both regions E. granulosus had the G1 genotype, and E. multilocularis was of the Asian type. Sequences alignment revealed 25 haplotypes for E. granulosus cox1 gene (15 in Xinyuan, 9 in Shiqu, and 1 shared by both) and 29 haplotypes for E. multilocularis (13 in Xinyuan, 15 in Shiqu, 1 shared by both). The haplotype networks of E. multilocularis and E. granulosus both showed a star-shaped network with a centrally placed main haplotype. There were 9 sequences of E. granulosus being the main haplotype type in Xinyuan (34.6%, 9/26), while only 1 sequence of that type was found in Shiqu (1/14), indicating a central role of Xinyuan County in the E. granulosus haplotype network. On the contrary, there were 23 sequences of E. multilocularis being the main haplotype type in Shiqu (58.9%, 23/39) and 7 of that type in Xinyuan (35.0%, 7/20), indicating that Shiqu County was more predominant in the E. multilocularis haplotype structure. Conclusion There is no apparent difference in the mitochondrial cox1 genotype of Echinococcus found between Xinyuan and Shiqu County, but there are some differences in the genetic polymorphism detected in the two sites. The genetic differentiation level of E. granulosus is higher than that of E. multilocularis over the same spatial span. These findings provide important referential basis for further study on transmission direction and dynamics of Echinococcus.

The effects of Echinococcus multilocularis protoscoleces infection on lymphocytes and their subpopulations in mouse liver and spleen

ZHANG Ning, ZHANG Chuan-shan, LI Zhi-de, LI Liang, WANG Hui, TAN Ya-chao, SHANG Lin-lin, WEN Hao

2020, 38(1):  30-35.  doi:10.12140/j.issn.1000-7423.2020.01.005

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Objective To investigate the effect of Echinococcus multilocularis protoscoleces infection on lymphocytes and their subpopulations in livers and spleens of C57BL/6 mice. Methods Thirty-two C57BL/6 mice were randomly assigned into four groups of eight animals each: control group, low-dosage group (50 protoscoleces/mouse), mid-dosage group (500 protoscoleces/mouse) and high-dosage group (2 000 protoscoleces/mouse). The mice received portal vein injection of 200 μl protoscolex suspension or normal saline. Four weeks after infection, four mice in each group were euthanized to collect liver tissues for histopathological observation with routine Masson staining, while the other four mice were euthanized for orbital blood collection and liver and spleen cell suspension preparation. The numbers of CD3⁺ T cells, B cells, NK cells, NKT cells, CD4⁺ T cells, CD8⁺ T cells, CD4⁺CD69⁺ T cells, and CD8⁺CD69⁺ T cells, as well as the percentages of naive CD4⁺ T cells (CD4⁺ Tn), effector memory CD4⁺ T cells (CD4⁺ Tem), naive CD8⁺ T cells (CD8⁺ Tn), effector memory CD8⁺ T cells (CD8⁺ Tem) and central memory CD8⁺ T cells (CD8⁺ Tcm) were detected by flow cytometry (FCM). The FCM data were analyzed with Flow-Jo software. Statistical analysis was performed using Graphad Prism 7.0 software. Results Four weeks after infection, there were no significant changes in the livers of the control mice. Both the low- and mid-dosage groups showed weakening of acute inflammatory cell infiltration in liver tissues, and some lesions had gradually transformed to the fibrotic repair stage, with formation of small granulomatous nodules and appearance of penctate or focal necrosis. In the high-dosage group, small vesicles were seen in the liver. Germinal layers were observed, accompanied by intensive inflammatory cell infiltration. The numbers of CD3⁺ T cells in mouse liver in the high-, mid- and low-dosage groups and the control group were (4.10 ± 0.04) × 10⁵, (3.81 ± 0.14) × 10⁵, (3.02 ± 0.12) × 10⁵, and (2.98 ± 0.03) × 10⁵, respectively, with that in the high-dosage group being significantly higher than the other 3 groups (P < 0.05 or 0.01). The numbers of CD3⁺ T cells in mouse spleen in the high-, mid- and low-dosage groups and the control group were (16.01 ± 0.40) × 10⁵, (11.03 ± 1.21) × 10⁵, (10.10 ± 1.01) × 10⁵, and (9.71 ± 0.90) × 10⁵, respectively (P < 0.01), with that in the high-dosage group being significantly higher than the other 3 groups (P < 0.01). The high-dosage group also had significantly higher numbers of CD4⁺ T cells and CD8⁺ T cells in livers as well as B cells and CD8⁺ T cells in spleens, compared with other groups (P < 0.05 or P < 0.01). The numbers of CD4⁺CD69⁺ T cells in mouse liver in the high-, mid- and low-dosage groups and the control group were (3.23 ± 0.10) × 10⁴, (1.98 ± 0.11) × 10⁴, (1.51 ± 0.26) × 10⁴, and (1.19 ± 0.21) × 10⁴, respectively, with that in the high-dosage group being significantly higher than the other groups (P < 0.05). The respective numbers of CD4⁺CD69⁺ T cells in mouse spleen in the high-, mid- and low-dosage groups and the control group were (10.10 ± 0.41) × 10⁴, (8.91 ± 0.80) × 10⁴, (8.20 ± 0.41) × 10⁴, and (6.81 ± 0.50) × 10⁴, with that in the high-dosage group being significantly higher than the other groups (P < 0.01). The high-dosage group also had significantly higher numbers of CD8⁺CD69⁺ T cells in livers and spleens, compared with other groups (P < 0.05 or P < 0.01). The percentages of CD4⁺ Tn in mouse liver in the high-, mid- and low-dosage groups and the control group were (15.52 ± 1.51)%, (19.3 ± 2.09)%, (20.66 ± 1.28)%, and (23.62 ± 2.84)% respectively, with that in the control group being significantly higher than the other groups (P < 0.05 or P < 0.01). The control group also had significantly higher percentages of CD8⁺ Tn cells in livers and spleens, compared with other groups (P < 0.05 or P < 0.01). The high-dosage group had significantly higher percentages of CD8⁺ Tcm cells in livers and spleens, compared with other groups (P < 0.05). Conclusion In C57BL/6 mice infected with high-dose E. multilocular protoscoleces, the liver T cell recruitment occurred mainly of CD4⁺ and CD8⁺ T cells, while B cell and CD8⁺ T cell were mainly recruited in spleen.

Prevalence and transmission characteristics of human echinococcosis in Lhasa area, China, in 2017

GESANG Zhuo-ga, TANG Xiao-jia, WANG Zheng-he, LI Ran, TAO Ran, WEN Bo

2020, 38(1):  36-40.  doi:10.12140/j.issn.1000-7423.2020.01.006

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Objective To understand the prevalence and characteristics of human echinococcosis in Lhasa City. Methods Population based screening for echinococcosis was performed in 2017 by abdominal ultrasound scanning using a portable ultrasound scanner. Diagnosis was made according to the Diagnostic Criteria for Echinococcosis (WS 257-2006), combined with epidemiological history and clinical manifestations. Suspected cases further received a serological test with ELISA to detect serum IgG antibody against Echinococcus antigen. Data were analyzed using the SPSS 20.0 statistical software. Results The overall detection rate of human echinococcosis in Lhasa City was 0.24% (935/392 592) in 2017. The detection rate in females (0.28%, 533/193 671) was significantly higher than that in males (0.20%, 402/198 921) (P < 0.05). Echinococcosis was found in all age groups, with the highest prevalence in the group of ≥ 70 years (0.90%, 106/11 769) and the lowest in the group < 10 years (0.09%, 48/54 181). In the age groups of 50-59, 60-69, and ≥ 70 years, the prevalence in females(0.51%, 111/21 812; 0.93%, 92/9 924; 1.06%, 70/6 629, respectively) was significantly higher than those in males (0.35%, 73/20 853; 0.57%, 51/8 988; 0.70%, 36/5 140, respectively) (P < 0.05). Among the groups of different educational levels, the illiteracy and semi-illiteracy group had the highest detection rate of echinococcosis (0.45%, 369/82 206), while the college and higher group had the lowest(0.14%, 26/18 124). In the illiteracy and semi-illiteracy and the primary school group, the detection rates in females (0.54%, 232/43 171; 0.36%, 148/40 655) were both significantly higher than those in males (0.35%, 137/39 035; 0.28%, 118/41 786) (P < 0.05). The occupation distribution of echinococcosis showed that the prevalence was found the highest in the herdsman-farmer group (1.20%, 59/4 923) and the lowest in students (0.12%, 79/64 398). The prevalence in females of herdsman group and the group of other occupations was 0.96% (93/9 675) and 0.26% (201/77 235), respectively, being significantly higher than those in males(0.52%, 53/10 139 and 0.14%, 113/79 483, respectively) (P < 0.05). The lifestyle distribution indicated that the prevalence was the highest in those settled in winter and nomadic in summer(1.70%, 26/1 529), among them the females showed the prevalence (2.65%, 21/792; 0.28%, 431/153 203) significantly higher than those in males(0.68%, 5/737; 0.20%, 309/156 215, respectively)(P < 0.05). Among the groups engaging in different types of production, the prevalence was found the highest in the group of livestock farming (0.57%, 128/22 351), and the lowest in the agriculture group (0.20%, 266/130 298). In all production type groups, except the herding-farming group, higher prevalence was found in females of all other groups including town residents (0.24%, 246/104 250), livestock farming group (0.73%, 82/11 162), and agriculture group (0.24%, 149/62 149) respectively, which were significantly higher than those in males (0.19%, 195/103 092; 0.41%, 46/11 189 and 0.17%, 117/68 149, respectively) (P < 0.05). Furthermore, the prevalence varied in different counties (districts), ranging at 0.18%-0.41%, with the highest in Damshung County(0.41%, 102/24 588) and lowest in Nyemo County (0.18%, 48/26 768). The prevalence was higher in the counties of Lhundup (0.28%, 135/48 848), Damshung (0.41%, 102/24 588) and Medro Gongkar (0.28%, 117/41 689), compared with Chengguan District (urban area of Lhasa) (0.21%, 378/180 594) (P < 0.05). Conclusion The survey indicates that females, people at age ≥ 70, in the occupation of herding-farming, with living style being settled in winter and nomadic in summer, illiterate and semi-illiterate, and engaging in livestock farming are at higher risk of contracting echinococcosis in Lhasa area.

Mitochondrial function regulated by Beclin1 in liver fibrosis in schistosomiasis

SHEN Shuang, LUO Jun-tao, YE Jian-ping

2020, 38(1):  41-46.  doi:10.12140/j.issn.1000-7423.2020.01.007

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Objective To explore the relationship between Beclin1 and mitochondrial function in hepatic fibrosis in schistosomiasis. Methods Forty female C57BL/6 mice were randomly divided into infected group and healthy control group (20 mice in each group). Mice in the infected group were infected with 20 Schistosoma japonicum cercariae by the abdominal patch method. Ten weeks after infection, fresh liver tissues were collected from the two groups. Liver pathology was assessed by HE and Masson staining of liver tissue sections. The relative protein levels of Beclin1, p-Beclin1 and voltage-dependent anion channel protein 1 (VDAC1) in liver were assessed by Western blotting. Fluorescence quantitative PCR was performed to analyze the relative mRNA expression of collagen Col 1a1 and Col 3a1, as well as enzymes involved in the mitochondrial citric acid cycle reaction, including citrate synthase, isocitrate dehydrogenase, alpha ketoglutarate dehydrogenase and malate dehydrogenase. Another part of liver tissues was sectioned for visualizing mitochondrion morphology by electron microscopy. To examine the mitochondria oxygen consumption rate (OCR) using cell energy metabolizer, the mitochondria extracted from liver tissues were added to the culture plates, to which complex V substrate ADP, complex V inhibitor oligomycin, uncoupler FCCP, and antimycin A (an inhibitor for complex Ⅱ plus Ⅲ) were sequentially added, allowing culture for 5 min, then the OCR was detected. Results HE and Masson staining showed that granulomas were formed in mouse liver in the infected group, and obvious fibrosis occurred around the granulomas. Western blotting showed that the protein level of VDAC1 remained unchanged in the infected liver, while Beclin1 (0.65 ± 0.05) and p-Beclin1 (0.78 ± 0.03) were decreased, compared with the control (P < 0.05). The fluorescence quantitative PCR results indicated that the relative transcription levels of Col 1a1 and Col 3a1 mRNA in livers of the infected group were 4.05 ± 0.23 and 2.71 ± 0.14, respectively, which were higher than those of the healthy control group (P < 0.01). The relative transcription levels of citrate synthase, isocitrate dehydrogenase, α-ketoglutarate dehydrogenase and malate dehydrogenase mRNA were 0.61 ± 0.03, 0.65 ± 0.01, 0.41 ± 0.03 and 0.55 ± 0.01, respectively, which were lower than those of the healthy control group (P < 0.01). Electron microscopy revealed that in the infected group, the mitochondrial structure was damaged and the ridge was broken. Mitochondrial function test showed that after addition of complex Ⅴ substrate ADP, the OCR of the healthy control group [(335 ± 29) pmol/min] was higher than that of the infected group [(78 ± 23) pmol/min] (P < 0.01); addition of complex Ⅴ inhibitor oligomycin reduced the difference [(80 ± 2) pmol/min in the control group and (31 ± 6) pmol/min in the infected group], although significance of difference still existed (P < 0.05); after addition of uncoupler FCCP, the OCRs of the healthy control and the infected groups increased to (159 ± 4) pmol/min and (42 ± 5) pmol/min(P < 0.01), respectively. After addition of complex Ⅱ plus Ⅲ inhibitor, the OCRs of both groups decreased to zero. Conclusion In the development of liver fibrosis in schistosomiasis, Beclin1 is reduced, the citric acid cycle is restrained, and the mitochondrial OCR further declines. The complex Ⅴ plays an important role in the regulation of mitochondrial activity during liver fibrosis in schistosomiasis.

iTRAQ-based proteomics of excretory-secretory products of Trichinella spiralis and Trichinella pseudospiralis at the muscle larva stage

ZHANG Yu-lu, WANG Yang, BAI Xue, TANG Bin, HU Xiao-xiang, ZHANG Chun-ling, LIU Ming-yuan, LIU Xiao-lei

2020, 38(1):  47-53.  doi:10.12140/j.issn.1000-7423.2020.01.008

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Objective To analyze differential proteins between excretory-secretory products (ESPs) of Trichinella spiralis and Trichinella pseudospiralis muscle larvae, and find out the causes for different immunosuppression between the two species and the potential functional proteins involved in the formation of cysts. Methods ESPs were collected from T. spiralis and T. pseudospiralis at the muscle larval stage. The bicinchoninic acid (BCA) detection method was used to determine the ESP protein concentrations. The quality of ESP proteins were assessed by sodium lauryl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), then the proteins were digested by filter-aided sample preparation (FASP), and labelled with isobaric tags for relative and absolute quantitation (iTRAQ) reagents. The samples with mixed labeling underwent reversed-phase high performance liquid chromatography (RP-HPLC) and liquid chromatography-tandem mass spectrometry (LC-MS/MS). Then the uniprot databases for T. spiralis uniport and T. pseudospiralis were searched, to screen for homologous proteins based on the trusted proteins (scores > 80) and compare their differential expression. Standardized description of differential proteins was done with QuickGO. Nine proteins with significant difference were selected for qRT-PCR verification using the ΔΔCt method. Statistical analysis was performed using the SPSS 19.0 software. Results Based on the iTRAQ analysis, we identified 492 credible proteins for T. spiralis, of which 193 were suitable for quantification, and 535 credible proteins for T. pseudospiralis, of which 164 could be quantified. T. spiralis/T. pseudospiralis homology comparison revealed that 162 proteins with up-regulation and 31 with down-regulation. The GO analysis results showed that the molecular functions were mainly involved in ion-binding function (45 proteins), peptidase activity(18), oxidoreductase activity(12), and nuclease activity (8). As regards biological processes, the small molecule metabolism processes involved the largest number of proteins (15), followed by the carbohydrate metabolism processes (12), the biosynthesis processes (12), and DNA metabolism processes (8). Of various cell components, the cytoplasm and protein-containing complexes had highest enrichment of differential proteins. KEGG analysis showed that most of the proteins related to thiamine metabolism, sphingolipid biosynthesis, and phenylalanine, tyrosine and tryptophan biosynthesis were up-regulated. In addition, at the muscle larva stage, E5SXW8 (plancitoxin-1), E5SPA7 (putative trypsin), E5S387 (cystatin), E5SJH4 (squash family serine protease inhibitor), E5S554 (5′-nucleotidase), E5SQX1 (enolase) and E5SBA6 (L-asparaginase) had higher protein and transcription levels in T. spiralis than in T. pseudospiralis (P < 0.05), while E5RZQ6 (heat shock protein beta-1) and E5S7K8 (histone H2B) had a reversed pattern (P < 0.05). Conclusion The bioinformatics analysis on differential excretory-secretory products between T. spiralis and T. pseudospiralis muscle larva stage suggests that serine protease, cystatin, plancitoxin-1 and 14-3-3 protein may involve in the formation of cysts and immunomodulation.

Molecular identification of Anopheles hyrcanus group and faunal distribution of Anopheles sinensis (Diptera ∶ Culicidae)in China

PENG Heng, CHEN Han-ming, CHEN Hui-ying, WANG Yan, LI Xiang-yu, LI Ji-xu, YANG Zhen-zhou, MA Ya-jun

2020, 38(1):  58-67.  doi:10.12140/j.issn.1000-7423.2020.01.009

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Objective To improve the multiplex PCR molecular methods to identify species of Anopheles hyrcanus group, and investigate the faunal distribution of An. sinensis in China and the factors influencing its distribution. Methods The species-specific primers were designed based on rDNA-ITS2 sequences of Anopheles (An. sinensis, An. lesteri, An. yatsushiroensis, An. kleini and An. belenrae) to improve the specificity and sensitivity of the multiplex PCR method to identify the Anopheles mosquito species. Anopheles mosquitoes were collected from 18 locations in 8 provinces (municipalities or autonomous regions) in China using light trap and entomological aspirator from 2013 to 2018. Those mosquitoes identified as members of An. hyrcanus group based on morphological characteristics were used for further analysis. The genomic DNA of individual mosquito was extracted, and the multiplex PCR assay was used to determine the species of An. hyrcanus group. For those mosquitoes without multiplex PCR products, the rDNA-ITS2 was amplified and sequenced, and the sequences were subjected to BLAST of the NCBI GenBank database to determine the species. Based on the results of this study and the review of 17 publications that identified An. sinensis using molecular biological methods in China, South Korea and the Far East of Russian, as well as the collected geographical location and climate data, the influence q value was calculated using the geo-detector software. The effects of geographical longitude, latitude, annual average temperature and rainfall on the distribution of An. sinensis were analyzed. Results The improved multiplex PCR method was able to simultaneously identify five members of the An. hyrcanus group based on the size of the amplified fragments as An. sinensis (490 bp), An. lesteri (313 bp), An. yatsushiroensis (216 bp), An. kleini (386 bp) and An. belenrae (165 bp). In this study, the species of 365 Anopheles mosquitoes were identified by multiplex PCR, including An. sinensis (n = 114, collected from Shaanxi, Anhui and Shandong), An. yatsushiroensis (n = 34), An. lesteri (n = 9), An. kleini (n = 181), An. belenrae (n = 5). The 22 individuals without acquiring amplified products were identified as An. kweiyangensis (n = 2), An. sineroides (n = 1), An. koreicus (n = 8), An. lindesayi (n = 7) and An. pattoni (n = 4) by rDNA-ITS2 sequencing. The geographic and ecological data were collected from 101 sites, including 80 sites with An. sinensis and 21 sites without An. sinensis distribution. The calculated q values were 0.592 0 for annual average temperature, 0.507 2 for latitude, 0.351 2 for longitude and 0.214 4 for annual average rainfall. The results indicate that the most significant factor affecting the distribution of An. sinensis is temperature, followed by latitude. Comprehensive analysis of the impact of latitude and annual average temperature on the distribution of Anopheles revealed that the annual average temperature of 10 ℃ could be used as the reference to determine the northern boundary of An. sinensis distribution in China. An. sinensis was distributed in the entire provinces (municipalities or autonomous regions) of Yunnan, Guizhou, Chongqing, Henan, Shandong, Tianjin, Jiangsu, Anhui, Hubei, Zhejiang, Shanghai, Fujian, Jiangxi, Guangxi, Guangdong, Hainan, Taiwan, Hong Kong and Macao, and the southern parts of the Tibet, Sichuan, Gansu, Shaanxi, Shanxi, Hebei, Beijing and Liaoning. Conclusion The improved multiplex PCR assay used to identify An. hyrcanus group is fast, simple and reliable. The annual average temperature of 10 ℃ could be used as reference to determine the northern boundary for the geographical distribution of An. sinensis in China, and the distribution range is smaller than that in the previously recorded document.

Polymorphisms of circumsporozoite protein gene and population structure analysis of Plasmodium vivax with different infection sources in Yunnan Province

XU Yan-chun, DONG Ying, DENG Yan, MAO Xiang-hua, CHEN Meng-ni, ZHANG Cang-lin, JIANG Lu-bin

2020, 38(1):  67-73.  doi:10.12140/j.issn.1000-7423.2020.01.010

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Objective To analyze the gene sequences of circumsporozoite protein of Plasmodium vivax (Pvcsp) collected from different infection sources in Yunnan Province, in order to understand the population structure and genetic diversity of Pvcsp gene in the Province. Methods Epidemiological data and blood samples were collected from the local and imported P. vivax malaria cases in Yunnan Province reported in the China Infectious Disease Network Direct Reporting System from 2014 to 2017. Genomic DNA of Plasmodium was extracted from blood samples, amplified by nested PCR and sequenced. MEGA 5.04 and Arlequin 3.5.2.2 were used to analyze the haploid type and the expected heterozygosity(He), respectively, and DnaSP 5.10 was used to calculate nucleotide diversity (π), synonymous replacement rate(Ks), missense replacement rate(Ka) and interpopulation genetic differentiation index (Fst). Results A total of 969 blood samples of P. vivax malaria cases were analyzed, and 759 samples were successfully amplified to produce target bands of 650-750 bp, including 39 cases in Yunnan, 16 cases from Africa, 688 cases from Myanmar, 13 cases from Laos, 2 cases from Cambodia and 1 case from Pakistan. The haploid type analysis showed the presence of 90 haploid types among the 759 Pvcsp gene sequences, of which 29 belonged to the PV-Ⅰ temperate family (similar to the VK210 type), 50 to the PV-Ⅰ tropical family (similar to the VK210 type), and 11 were of the PV-Ⅱ type (similar to the VK247 type). The proportions of three genotypes to the total examinees were 51.4% (390/759), 41.1% (312/759), and 7.5% (57/759), respectively, distributed in Yunnan local and Myanmar imported cases, while imported cases from Africa, Laos and other places only exhibited the PV-Ⅰ type. The amino acid sequence mutations of the PV-Ⅰ and PV-Ⅱ types occurred at 29 and 10 loci, respectively. The He, π and Ka/Ks of the 759 samples were 0.224, 0.075, and 0.480, respectively. Among the infection sources other than Cambodia/Pakistan, Pvcsp had the highest genetic diversity within imported cases from Laos(He, 0.422; π, 0.03). Pvcsp in Yunnan local cases and the imported cases from Africa showed the highest genetic differentiation (Fst, 0.082), while the lowest genetic differentiation was seen in the imported cases from Myanmar (Fst, 0.002); the Pvcsp gene displayed moderate genetic differentiation in the imported cases from Africa and Southeast Asia; and minor genetic differentiation was deteted in the imported cases from Southeast Asia. Conclusion There are three genotypes of Pvcsp of P. vivax from different sources of infection detected in Yunnan Province, with PV-Ⅰ temperate family as the dominant strain. The population genetic structure and differentiation varies with different genptypes.

Specific immunotherapeutic effect of recombinant Der p1 T cell epitope fusion protein on asthma in a mouse model

ZHAO Ya-nan, HONG Yong, LI Chao-pin

2020, 38(1):  74-79.  doi:10.12140/j.issn.1000-7423.2020.01.011

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Objective To explore the specific immunotherapeutic effect of Der p1 T cell epitope fusion protein on asthma in a mouse model. Methods Total 30 female BALB/c mice were randomly divided into three groups: control group, asthma group and specific immunotherapy group(SIT) treated with Der p1 T epitope fusion protein. The mice in asthma and SIT groups were sensitized with intraperitoneal injection of 200 μl(10 μg) crude extracts of house dust mite on day 0, 7 and 14, while the mice in control group were given with intraperitoneal injection of the same volume of PBS solution. Then the mice in asthma and SIT groups were challenged by spray inhalation of house dust mite extracts (0.5 μg/ml) for 30 minutes every day started from 21st day. The asthmatic symptoms of the challenged mice were observed and recorded for 7 consecutive days. Control group was given with the same volume of PBS. The mice in SIT group were treated with intraperitoneal injection of 20 μg Der p1 T cell epitope fusion protein in total volume of 200 μl 30 min before each challenge for 7 consecutive days. Mice in control group and asthma group were injected intraperitoneally with the same volume of PBS. The blood was collected from each mouse and sera were collected 24 hours after the last challenge, mice were euthanized and bronchoalveolar lavage fluid was collected by trachael intubation. ELISA kit was used to detect the serum levels of antigen-specific IgE and IgG2a and cytokines, including IL-10, IL-4, IFN-γ and IL-17A in the bronchoalveolar lavage fluid. Flow cytometry analysis was used to determine the change of Th1/Th2 and Th17/Treg cells. Lung tissues were taken for histological observation of the pathological changes. Results After being sensitized and challenged, the mice in the control group only had a short period of mild restlessness. The mice in asthma group showed obvious symptoms of restlessness, wheezing and out of breath. After immunotherapy, the mice in the SIT group had reduced asthmatic symptoms, significant improvement than mice in asthma group without treatment. The results of ELISA showed that the amount of specific IgE antibody in asthma group was (31.49 ± 4.32) IU/ml, which was significantly higher than that in control group [(8.53 ± 1.92) IU/ml] (P < 0.01) and in SIT group [(16.68 ± 2.45) IU/ml] (P < 0.01). The amount of antibody IgG2a in asthma mice was (19.56 ± 3.89) μg/ml, which was lower than that in control group [(42.43 ± 2.07) μg/ml] (P < 0.01) and in SIT group [(36.96 ± 5.04) μg/ml] (P < 0.01). The levels of IFN-γ and IL-10 in asthma group were (134.23 ± 22.49) pg/ml, (22.43 ± 8.27) pg/ml, respectively, which are significantly lower than those in control group [(212.36 ± 33.21) and (72.84 ± 21.42) pg/ml, respectively] (P < 0.05, 0.01). However, after being treated with SIT, the levels of IFN-γ and IL-10 had significantly recovered compared to the asthma group without SIT treatment, as (183.76 ± 24.66) pg/ml, (61.05 ± 7.97) pg/ml, respectively (P < 0.05, 0.01). The levels of IL-4 and IL-17A in asthma group were (165.45 ± 34.59) pg/ml, (464.21 ± 41.36) pg/ml, respectively, which are significantly higher than those in control group [(21.21 ± 5.26) and (115.74 ± 30.82) pg/ml, respectively] (P < 0.05, 0.01). Treatment with SIT reduced IL-4 and IL-17A levels to (64.15 ± 17.33) pg/ml, (271.61 ± 27.07) pg/ml, respectively, with significant difference compared to that in asthmat group without treatment (P < 0.05, 0.01). Consistently, the results of flow cytometry showed that the percentages of Th1 and Treg cells in CD4⁺ T lymphocytes of spleen in asthma group were 2.8% and 4.9% respectively, which were significantly lower than those in control group (3.7% and 10.3%) (P < 0.05, 0.01). The treatment with SIT significantly stimulated Th1 and Treg cells to 3.1% and 8.8%, respectively, with statistical significance compared to asthma group without treatment (P < 0.05, 0.01). The percentages of Th2 and Th17 cells in CD4⁺ T lymphocytes of spleen in asthma group were 2.8% and 2.2% respectively, which was significantly higher than those in control group (1.0% and 0.3%) (P < 0.01). The treatment of SIT reduced Th2 cells and Th17 cells to 1.7% and 0.6% respectively, with significant difference compared to asthma group without treatment (P < 0.01). Pathological results showed that there was significant inflammatory cell infiltration around the bronchioles, broken muscle fiber and the collapsed epithelial cells were observed in asthma group. These pathological changes were much improved in mice with SIT treatment with reduced bronchus wall thickness and inflammatory cells infiltration. Conclusion Der p1 T epitope fusion protein has significant immunotherapeutic effect on asthma in a mouse model.

Detection of schistosomiasis transmission risks in Yunnan Province based on ecological niche modeling

HU Xiao-kang, HAO Yu-wan, XIA Shang, GUO Yun-hai, XUE Jing-bo, ZHANG Yun, WANG Li-fang, DONG Yi, XU Jing, LI Shi-zhu

2020, 38(1):  80-87.  doi:10.12140/j.issn.1000-7423.2020.01.012

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Objective To predict the transmission risks of schistosomiasis based on ecological niche modeling and identify high-risk areas, in order to provide scientific evidence for the formulation of monitoring and control measures in Yunnan Province. Methods Village-level schistosomiasis epidemic data and 13 environmental variables such as climatic, geographical and socioeconomic factors were collected from 18 endemic counties in Yunnan Province from 2004 to 2015. BIOCLIM, DOMAIN and MaxEnt models were used to predict the schistosomiasis transmission risks in Yunnan Province, and the accuracy of prediction was assessed with the receiver operating characteristic area under curve (AUC). The model with best performance was used to analyze the importance of environmental variables and predict the distributions of schistosomiasis transmission risks in Yunnan Province. Results All the three models had good performance in predicting the distributions of schistosomiasis transmission risks in Yunnan Province, with the MaxEnt model having the highest prediction accuracy (AUC, 0.96 ± 0.01), followed by DOMAIN (AUC, 0.93 ± 0.04) and BIOCLIM (AUC, 0.88 ± 0.01) (P < 0.05 among three). The MaxEnt model revealed the annual average precipitation as the most significant environmental factor influencing the distributions of schistosomiasis (contribution value, 1.52), followed by gross domestic product and population density (contribution values 1.06 and 1.03, respectively). As predicted by the MaxEnt model, the transmission risk area, which was located mainly in the northwest, accounted for 3.1% of the area of Yunnan Province, comprising 2.7% of middle- and low-risk areas and 0.4% of high-risk areas. The high-risk areas were mainly distributed in northern Heqing County, eastern Eryuan County, central Dali City, northeastern Weishan County and northern Midu County. Conclusion It is feasible to predict distributions of schistosomiasis transmission risks based on the MaxEnt model. There still remain risks of schistosomiasis transmission in Yunnan Province, and the distributions of high-risk regions show a pattern of clustering. Therefore, targeted monitoring and control is needed.

Sequence analysis of ribosomal ITS2 gene and mitochondrial CO1 gene of Paragonimus metacercariae from freshwater crabs in Henan, Anhui, Fujian and Zhejiang provinces, China

HU Kun-min, CHEN Shao-hong, AI Lin, ZHENG Bin

2020, 38(1):  87-94.  doi:10.12140/j.issn.1000-7423.2020.01.013

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Objective To analyze the sequences of ribosomal second internal transcribed spacer（ITS2） and mitochondrial cytochrome c oxidase subunit Ⅰ (CO1) genes of Paragonimus metacercariae in 5 areas from Henan, Anhui, Fujian and Zhejiang Provinces of China, and identify the species of Paragonimus metacercariae. Methods From October 2018 to September 2019, freshwater crabs were collected from five survey sites, Lantian Town in Xiuning County of Anhui Province, Yongjia County in Zhejiang Province, Shaoyuan Town in Jiyuan City of Henan Province, as well as Zhenghe County and Zhangzhou City in Fujian Province. Genomic DNA was extracted from Paragonimus metacercariae from the crabs, and the ribosomal ITS2 and mitochondrial CO1 genes were amplified by PCR and sequenced. The amplified sequences were spliced and aligned by DNA Star software along with Blast alignment with the Paragonimus genes in GenBank^TM. Based on the ITS2 and CO1 gene sequences, a phylogenetic tree was constructed with the neighbor-joining method using Fasciola hepatica as the outgroup. Results The metacercaria positive rates found in the crabs from the five survey sites in Lantian Town, Xiuning County (Anhui Province), Yongjia County (Zhejiang Province), Shaoyuan Town, Jiyuan City (Henan Province), Zhenghe County and Zhangzhou City (Fujian Province) were 82% (49/60), 41% (29/70), 55% (41/74), 65% (41/63), and 45% (32/71), respectively. The lengths of amplified ITS2 and CO1 gene sequences were approximately 500 bp and 450 bp, respectively. The ITS2 and CO1 gene sequences of Paragonimus metacercariae from Lantian Town of Xuning County(Anhui Province) and Yongjia County (Zhejiang Province) showed the highest similarity with those of P. westermani (KC417492.1 and AF219379.2), being 99%-100% and 96%-99%, respectively, and both clustered in the same branch with P. westermani. The ITS2 and CO1 genes of Paragonimus metacercariae from Shaoyuan Town of Jiyuan City(Henan Province) showed the highest similarity with those of P. skrjabini (KX129924.1 and MK568551.1), at 98%-100% and 95%-99%, respectively, both being clustered in the same branch with P. skrjabini. The ITS2 gene of Paragonimus metacercariae from Zhenghe County(Fujian Province) showed the highest similarity (98%-100%) with that of P. skrjabini (KX129924.1), forming a clade with P. skrjabini and P. miyazaki (the distance between the two species was not obvious). The CO1 gene from Zhenghe County showed the highest similarity (91%-94%) with that of P. miyazaki (AY618823.1 and AY618834.1), formed a clade with P. miyazaki but was distant from the clade of P. skrjabini. The ITS2 gene of Paragonimus metacercariae from Zhangzhou City (Fujian Province) showed the highest similarity (100%) with the gene of P. westermani (KC417492.1), and the CO1 gene sequence showed the highest similarity (97%-99%) with that of P. cenococopiosus (AF159595.1), being clustered in the with P. cenococopiosus branch. Conclusion The CO1 gene sequence amplified from the metacercariae of 5 areas in 4 Provinces was highly homologous to those of P. westermani, P. skrjabini, P. miyazaki and P. cenococopiosus. The CO1 gene of Paragonimus may be used as a potential molecular marker for species differentiation.

Morphological and molecular characterization of mite Tyrophagus putrescentiae

XU Wei, ZHU Zhi-wei, LUO Xin, SUN En-tao, ZHAO Jin-hong

2020, 38(1):  95-101.  doi:10.12140/j.issn.1000-7423.2020.01.014

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Objective To identify the morphological and molecular characterization of mite Tyrophagus putrescentiae. Methods T. putrescentiae mites were collected from the flour stored over 1 year and the life cycle was maintained in the lab. The larval, nymph and adult mites and the eggs were observed under optical microscope and scanning electron microscope (SEM). The DNA was extracted from the mites, the cytochrome oxidase subunit Ⅰ(cox1) and internal transcribed spacer(ITS) genes were amplified by PCR. The PCR products were ligated into the vector pMD19-T, then transformed into Escherichia coli. The inserts in the positive clones were DNA sequenced. The obtained DNA sequences were analyzed by DNAStar software and performed BLAST alignment analysis against mite sequences in GenBank. The phylogenetic tree was constructed accordingly. Results The body of T. putrescentiae was oval shaped with a smooth, colorless and translucent surface. The pincers and feet were brownish, and the setae on the body were slender but not stiff. Morphological structures of T. putrescentiae, including legs, setae, external genitalia and accessories, could be clearly identified under SEM. The larva had three pairs of legs, the nymph and adult had four pairs of legs and one pair of chela. The body was composed of three segments. Within Ⅳ coxa, the reproductive organ can be discovered, the lateral plates bended outward to form a "herringbone" shape, and the anus was slender and located in the posterior half with longer anal hair beyond the body. The hair d1 is short, the d2 is longer, and hair hi is longer than the hair he. The clearly discovered hair pa in the posterior is beyond the body. The hair vi and outer hair ve were found in the forefoot, the hair sci was longer than that of the sce, and the outer hair o1 of the knee was slightly longer than that of o2. The PCR products of ITS amplified from T. putrescentiae was 1 852 bp and cox1 was 469 bp in length. BLAST search confirmed that cox1 sequence matched the sequence of T. putrescentiae. The A+T content of cox1 sequence was 65.7%. The phylogenetic tree based on cox1 gene sequence showed that the T. putrescentiae is genetically close to T. longier with sequence similarity of 91.5%. The A+T content of ITS sequences was 53.6%. The phylogenetic tree based on ITS showed that the sequence of ITS of T. putrescentiae was quite different from that of other Acarida, with the sequence similarity of 37.7%-41.8%. Conclusion The mites collected from the stale flour were identified as T. putrescentiae based on their morphological characterization and amplified cox1 and ITS gene sequences.

Investigation on the infection of Paragonimus skrjabini in intermediate hosts in northern Fujian

CAI Mao-rong, LUO Jun, AI Lin, LIAO Qi, ZHUO Ming-yin, CHEN Jia, CHEN Jin-zhong, LIU Jian-fa, CHENG You-zhu

2020, 38(1):  102-109.  doi:10.12140/j.issn.1000-7423.2020.01.015

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Objective To investigate the species and habitats of snails and crabs related to the infection of Paragonimus skrjabini in northern Fujian, and the association of environment changes to the infection of the parasite in these intermediate hosts. Methods Based on the living areas of P. skrjabini infected cases, the snails and crabs were collected from the areas in 11 sites in northern Fujian. The ecological environment of these sites, distribution density of snails and crabs and their infection with Paragonimus cercariae(snail) and metacercariae(crab) were investigated. The metacercariaes collected from infected crabs were used to infect two dogs. The specific internal transcribed spacer 2 (ITS2) and cytochrome oxidase 1(CO1) genes were amplified from the collected metacercariaes by PCR. The effect of environment changes in these habitat areas on the infection of P. skrjabini was investigated. Results Psedobhythinella tangi, P. jianouensis and Tricula xiaoqiaoensis were found in all 11 survey sites. These sails lived in the water ditches or wetlands under the mountain, mostly inhabited in the humid environment in the ditches 5 mm above and below the water line. Snails adhered to defoliated leaves or dead wood branches in ditches (62.63%), on stones (35.07%) and in mud (2.29%). The density distribution of P. tangi, P. jianouensis and T. xiaoqiaoensis was 156, 179-291 and 12-266 per cage, respectively. P. skrjabini cercariae were identified in all 11 survey sites with infection rate of 0.22% (4/1 851) in P. tangi, 0.38% (36/9 420) in P. jianouensis and 1.10% (102/9 247) in T. xiaoqiaoensis. Four species of crab were identified in these sites including Sinopotamon fukinense, Bottapotamon fukinense, Huananpotamon angulatum, and Huananpotamon spp. The infection rate of P. skrjabini metacercariae in these crabs in 11 sites varied from 38.99% to 96.77% with average infection rate of 80.21% (231/288) and infection intensity of 19.8 metacercariae per crab. PCR results showed that a fragment of 500 bp ITS2 DNA was amplified from collected metacercariaes that shares 99% sequence identity with the same gene from P. skrjabini deposited in GenBank, and 450 bp CO1 that shares 100% sequence identity with P. skrjabini CO1. The collected metacercariaes were used to infect two dogs and 6 and 8 adult worms were harvested from each infected dogs, all identified as P. skrjabini. The environment changes including flooding, plantation, droughts damaged the habitats of snail and crab and affected their infections of P. skrjabini with statistical difference (P < 0.05). No snail was found in 7 investigations and no crab was found in 3 sites after flooding. Conclusion The mountain areas in northern Fujian including Jianou are the natural endemic areas for the infection of P. skrjabini. The environment change including flooding and plantation are the main factors affecting the breeding of intermediate host snails and crabs and the infection of P. skrjabini in these areas.

INFORMATION EXCHANGES

A survey among health professionals working on tropical disease control on knowledge of training needs for global health

LI Hong-mei, DING Wei, HUANG Lu-lu, MA Xue-jiao, DUAN Lei, QIAN Ying-jun, WANG Duo-quan, GUAN Ya-yi, XIAO Ning, ZHOU Xiao-nong

2020, 38(1):  110-114.  doi:10.12140/j.issn.1000-7423.2020.01.016

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A survey was made on attendees of three conferences (or training courses) on tropical disease prevention and control held by the National Institute of Parasitic Disease, China CDC from August 2016 to April 2017, by using questionnaires to collect information. Data were analyzed using SPSS 20.0 software. A total of 256 questionnaires were distributed and 241 were valid. Among the respondents, 14.5% (35/241) had never heard of the term “global health”, 82.6% (199/241) had heard of it but did not understand the concept, and only 2.9% (7/241) were familiar with global health. The participants were significantly more familiar with epidemiology and health statistics than global health. There were 42.7% (103/241) respondents having correct understanding of the basic concept of global health, among them, the age group ≤35 had significantly higher percentage of correct understanding (53.8%, 56/104) than that in the group of age > 35 (34.3%, 47/137). In addition, 97.1% (233/240) of the respondents showed willingness to receive global health training. The top three demanded training contents were disease prevention and control strategies and techniques, technologies on experimental testing, diagnosis, research and development, and knowledge of health management. The top three demands for capacity improvement were global health vision, disease prevention and control experiences, and research and analysis capabilities. Trainings should be carried out primarily by on-site teaching, in an interactive teaching mode between trainers and trainees, inviting more teachers from foreign health systems. The extent of awareness on global health knowledge is challenging in the health personnel working in the prevention and control of tropical diseases in China, thus, it is urged to strengthen the training on global health to improve the capacity building.

REVIEWS

Current applications of metal nanoparticles in detection and treatment of common infectious diseases

JI Ya-nan, LIAN Xiao-li, ZHU Feng-long, YANG Yi-mei

2020, 38(1):  115-119.  doi:10.12140/j.issn.1000-7423.2020.01.017

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With the rapid development of nanotechnology in recent years, metal nanoparticles have shown promising future in application in medicine, and being gradually popularized due to their unique physicochemical properties. They are advantageous over traditional methods in detection and treatment of common infectious diseases. This review introduces applications of nanogold-based immunolabeling technology in pathogen detection, mainly involving in immunochromatography, biosensor preparation, and probe technology, as well as updated research advances in application of nanogold and nanosilver in anti-infection treatment.

Involvement of cysteine proteases in Acanthamoeba encystment

LI Qing-shan, WANG Zhi-xin, CHENG Xun-jia

2020, 38(1):  120-123.  doi:10.12140/j.issn.1000-7423.2020.01.018

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Acanthamoeba is an opportunistic protozoan widely distributed in the environment, and can cause severe and even life-threatening infections in humans. Acanthamoeba undergoes two stages during the life cycle: the pathogenic trophozoite stage and the infectious cyst stage. Further, there are two stages during encystment, including the early stage involving autolysis and protein degradation and the late stage with cyst-specific gene expression. Cysteine proteases play an important role in the early stage of encystment, which could be inhibited by the cysteine protease inhibitor E64d. Cysteine proteases are cathepsin L-like enzymes with a conserved catalytic site, expressed in trophozoites and cysts continuously, and are localized in lysosomes, exhibiting highest catalytic activities under acidic condition. Unlike serine proteases, cysteine proteases extensively hydrolyze proteins during the encystment process. The cystic-specific cysteine proteases play a key role in encystment of Acanthamoeba through mediating mitochondrial autophagy, this paper reviews the effects of endogenous cysteine protease inhibitors modulating the encystment process.

Research progress on the correlations of tick-borne diseases with meteorological factors and their prevention measures in China

YAO Qing-mei, ZHOU Su-fang, ZHANG Yi, XIA Shang, XUE Jing-bo

2020, 38(1):  123-127.  doi:10.12140/j.issn.1000-7423.2020.01.019

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Ticks can carry pathogens that may transmit a variety of zoonotic diseases while feeding on the blood of animals. The tick-transmitted diseases, collectively referred to as tick-borne diseases, are correlated with meteorological factors, particularly temperature, relative humidity, rainfall, relative atmospheric pressure and wind speed. This paper gives an overview on the progress in research on the correlations of tick-borne diseases with meteorological factors and their prevention measures in China, in order to set up a meteorological model for tick-borne diseases, and establish meteorological early warning and forecasting systems for predicting the occurrence and monitoring the development of the diseases.

Research advances on the relationship between cestode excretory/secretory products and host immune response

FAN Xian-min, ZHOU Bi-ying

2020, 38(1):  128-133.  doi:10.12140/j.issn.1000-7423.2020.01.020

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Cestodes are common zoonotic parasites. The excretory/secretory products (ESPs) of cestodes are produced during the infection and exposed directly to the host immune system, being one of the most possible effectors causing the changes of host immune responses. In recent years, a wealth of studies have shown that ESPs can induce and regulate the host immune responses, which are closely related to the cestodes infection. Cestodes ESPs can induce host’s positive immune response to control the infection, and also induce negative immune response, enabling the cestodes to evade the host immune attacks. thus resulting in establishment of a long-time infection. This paper presents a brief review on the relations between cestode ESPs and host immune response.

SHORT COMMUNICATIONS

Analysis of hospital expenses for patients with end-stage hepatic alveolar echinococcosis receiving ex vivo liver resection and autotransplantation

KASIMU Aihaiti, ABUDUSALAMU Aini, TUERGANAILI Aji, SHAO Ying-mei, ZHANG Rui-qing, TALAITI Tuergan, JIANG Tie-min, RAN Bo, ABUDUAINI Abulizi, MIERADILI Aierken, WEN Hao

2020, 38(1):  53-57.  doi:10.12140/j.issn.1000-7423.2020.01.021

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We retrospectively collected demographic information as well as relevant clinical data of patients with end-stage hepatic alveolar echinococcosis (AE) treated with ex vivo liver resection and autotransplantation (ELRA) in the First Affiliated Hospital of Xinjiang Medical University during 2010-2018. The patients were divided into six groups based on their medical history prior to ELRA: group A patients having no relevant disease history record; group B having anti-parasitic chemotherapy history; group C having the history of anti-parasitic chemotherapy plus hepatoprotective medication; group D having the record of alleviating jaundice by percutaneous or endoscopic approach plus hepatoprotective medication; group E experienced interventional treatment via liver blood vessel plus anti-parasitic chemotherapy; and group F experienced hepatectomy and anti-parasitic chemotherapy. The medical expenses of different groups were compared by the Mann-Whinty test. A total of 82 patients receiving ELRA were included in this study, with a male/female ratio of 39 : 43 and an average age of (36.12 ± 11.84) years. Most of them were of Zang, Kazakh and Han ethnity, and came from Xinjiang, Tibet and Sichuan. The medical expenses for the hepatic AE patients receiving ELRA treatment included the costs of western medicine, medical examinations, consumable material, ELRA surgery and special material. The average total expense of the 82 patients was (241 155.0 ± 119 746.5) yuan, of which the cost of western medicine accounting the highest proportion (41.1%), followed by the cost of medical examinations (15.7%), consumable material (13.4%), surgery (9.6%) and special material (7.5%). The groups C and D had significantly higher total expenses (P < 0.01) than the other 4 groups. When compared with itemized cost, the group C and D expensed significantly higher for western medicine, medical examinations, nursing, and treatment (P < 0.05), compared with other 4 groups. The group C and D also expensed significantly higher for special material, blood supply, emergency aid, and ward bed than those of group A (P < 0.05). The total and itemized expenses of the patients with no relevant disease history prior to surgery were lower than those with these history records. In addition, occurrence of jaundice or liver dysfunction may increase the total expense and the costs of most items.