After more than 70 years of effective control programme, China has made remarkable achievements in the control of key parasitic diseases, and is moving towards the goal of control and elimination. This paper analyzes the epidemic status and challenges of schistosomiasis, malaria, echinococcosis, leishmaniasis, clonorchiasis and soil-transmitted helminthiasis in recent years, and puts forward the future directions and key tasks of those diseases, in order to provide reference for accelerating the control and elimination programmes on key parasitic diseases in China.
Objective To understand the situation of soil-transmitted nematode infection in China in 2020 and provide support for evaluating the development of surveillance on soil-transmitted nematodiasis in various provinces/municipalities/autonomous regions, and improving and perfecting the control strategies. Methods Surveillance was carried out in 408 national surveillance sites (counties) in 31 provinces (municipalities, autonomous regions) in China in 2020. With the county as unit, each site was divided into 5 areas geographically: east, west, south, north, and central part, followed by selecting one township (town), and therein one administrative village was selected from wherein, 200 permanent residents over the age of 3 were sampled. A total of 1 000 people were surveyed at each surveillance site. Fecal samples were collected from the sampled villagers, and examined by using the modified Kato-Katz thick smear method (two slide-reading for each sample) for infection of hookworm, Ascaris lumbricoides, Trichuris trichiura and Enterobius vermicularis, to calculate the infection rate and intensity, respectively. In addition, soil samples were collected from fields or vegetable gardens of each village in the survey site, and examined for hookworm larvae using 5% saline at 45 ℃, and for Ascaris eggs by saturated sodium nitrate flotation method. Results In 2020, the overall infection rate of soil-transmitted nematode in residents was 0.84% (3 485/415 672) in 408 surveillance sites of 31 provinces (municipalities/autonomous regions), with the highest found in Hainan (6.34%, 199/3 141), followed by Yunnan (5.80%, 963/16 616) and Sichuan (3.66%, 592/16 168); infection rate in females was 0.91% (1 944/213 591), which was higher than that of 0.76% in males (1 541/202 081) (χ2 = 27.20, P < 0.01). The soil-transmitted nematode infection rate was the highest in the age group ≥ 60-years-old, which is 1.26% (1 376/109 251). The difference between each age group was statistically significant (χ2 = 382.28, P < 0.01). The infection rates of hookworm, A. lumbricoides, T. trichiura were 0.51% (2 016/415 672), 0.19% (805/415 672) and 0.16% (673/415 672), respectively. Among them, hookworm and T. trichiura had only mildly infected cases. The proportions of mild and moderate A. lumbricoides infections were 99.25% (799/805) and 0.75% (6/805), respectively. In 2020, 2 604 soil samples were examined and found that the positive rate of Ascaris eggs and hookworms was 3.07% (80/2 604) and 2.42% (63/2 604), respectively. Conclusion In 2020, the infection rate of soil-transmitted nematode in China remains at a low level in general, but the regional differences are still significant, and the areas with high infection rates still exist. At the same time, it is necessary to strengthen the control measures for the key groups of people over age of 60, women and children, and carry out health education.
Toxoplasma gondii is an obligate intracellular parasite that widely distributes in the world and causes zoonotic toxoplasmosis. In recent years, parasite infection in the brain has been paid more and more attention. T. gondii can cause central nervous system damage, often manifested as depression, schizophrenia, epilepsy and other symptoms. In this paper, the process and mechanism of T. gondii establishing chronic infection through blood-brain barrier, causing central nervous system injury and disease are reviewed.
To understand the Anisakis larvae infection status in marine fishes sold in Shanghai, the fresh marine fishes caught in the East China Sea area were collected from Farmers’ markets, supermarkets and seafood markets in Shanghai in 2022. The suspected Anisakis larvae were searched in the offal and muscles after dissection and observed under optical microscope and scanning electron microscope, respectively. A total of 338 marine fish of 16 species were collected, and 1 065 Anisakis larvae were found from 116 fish of 6 species, with a total infection rate of 34.3% (116/338) and average infection intensity of 9.2 larvae/fish. The highest infection rate was 11/12 in Lophiiformes, and the highest average infection intensity was 13.0 larvae/fish in Larimichthys polyactis. The Anisakis larvae infection rates increased gradually from spring to winter. The infection rate and average infection intensity in winter were 51.1% (46/90) and 12.3 larvae/fish, respectively, which were the highest seasons of the year. The predominant sites of Anisakis larvae parasitise in marine fishes were the intestines and abdominal cavity, with infection rates of 54.6% (582/1 065) and 40.7% (433/1 065), respectively. The result showed that Anisakis larvae infections were present in marine fish sold in Shanghai and the infection rates of commonly consumed marine fishes such as Lophiiformes and L. polyactis were high.
Food-borne parasitic diseases caused by ingesting food and water containing infective parasites are still common parasitic diseases that are easily misdiagnosed and mistreated in clinical practice. With the participation of multi-disciplinary experts, and in the light of the latest research results at home and abroad, based on factors other than the quality of evidence (economics, patient preferences and values, trade-offs, accessibility, fairness, acceptability, etc.), the level of recommendation and the quality of evidence in evidence-based medicine were assessed using the World Health Organization-recommended evidence quality classification and strength of recommendation system, and a consensus of 24 items was reached to guide and improve the comprehensive diagnosis and treatment of food-borne parasitic diseases for clinical medical staff.
Objective To screen the differentially expressed genes (DEGs) by comparing the transcriptome of the brain tissues between the mice chronically infected with Toxoplasma gondii and normal mice, to analyze the relative transcription level of DEGs in the depression-related kynurenine (KYN) pathway and to provide a theoretical basis for exploring the mechanism of depression-like symptoms caused by Toxoplasma gondii chronic infecttion in mice. Methods SV129 male mice (n = 18) were randomly and equally divided into the infection group and the control group. Mice in the infection group were intraperitoneally injected with 120 tachyzoites of T. gondii ME49 strain (200 μl), and mice in the control group were injected with the same volume of PBS. After 3 months post-infection, mice brain tissues of the two groups were collected for extraction of total RNA to undertake transcriptome sequencing for screening DEG. With the DEGs obtained, cluster analysis, gene ontology (GO) functional annotation analysis, Kyoto Encyclopedia of Genes and Genomes (KEGG) functional annotation and enrichment analysis were performed. Eight DEGs [interferon-γ (IFN-γ), indoleamine 2,3-dioxygenase 1 (IDO1), IDO2, kynurenine-3-monooxygenase (KYNU), kynurenine-3-monooxygenase (KMO), 3-hydroxyanthranilate 3,4-dioxygenase (3-HAO), vimentin (Vim) and brain-derived neurotrophic factor (BDNF)] related to KYN pathway associated with depression were selected to examine each gene’s relative transcription level by quantitative real-time PCR (qRT-PCR), using glyceraldehyde-3-phosphate dehydrogenase gene as an internal reference. Results Transcriptome sequencing found 2 295 DEGs in the brain of the mice from the infection and control groups, of which 2 016 were up-regulated and 279 were down-regulated. GO analysis showed that localisation was the most significantly enriched biological process, with a total of 257 DEGs. The most significantly enriched in cellular components was the protein-containing complex, with a total of 425 DEGs. The most significantly enriched molecular function was molecular transducer activity, with 177 DEGs. The largest number of DEGs enriched in biological process, cell component and molecular function were cell process, cell part and binding, with 1 039, 1 240 and 1 088 DEGs, respectively. KEGG analysis showed that the top three up-regulated metabolic pathways were the immune system, signaling transduction, and viral infectious disease, and the top three down-regulated pathways were signal transduction, signaling molecules and interaction and immune system. Functional enrichment analysis showed that 77 pathways were significantly enriched. The signaling pathways related to depression include tumor necrosis factor signaling pathway, neuroactive ligand-receptor interaction, NF-kappa B signaling pathway, JAK-STAT signaling pathway, necroptosis, apoptosis, chemokine signaling pathway, KYN pathway. The qRT-PCR results showed that the relative transcription levels of IFN-γ, IDO1, IDO2, KYNU, KMO, 3-HAO and Vim genes in the infection group were 3 023.08%, 355.52%, 190.17%, 496.55%, 339.92%, 212.74% and 507.34%, if the relative transcript level of control mice was taken as 100%. Compared with the control group, the transcription was significantly up-regulated (t = 3.782, 3.749, 3.226, 2.908, 2.533, 5.656, 2.948; all P < 0.05 or 0.01). The relative transcription level of BDNF was 63.32%, which was significantly down-regulated (t = 2.398, P < 0.05). The fold change of IFN-γ, IDO1, IDO2, KYNU, KMO, 3-HAO, BDNF, Vim obtained by qRT-PCR was 4.96, 1.74, 0.89, 2.10, 1.60, 1.06, -0.94, 2.18, respectively. The fold change obtained by transcriptome sequencing was 7.30, 0.55, 0.80, 3.83, 2.75, 3.53, -0.86 and 1.93, respectively. The transcriptional trend obtained by qRT-PCR was consistent with that obtained by transcriptome sequencing. Conclusion DEGs from brain tissues of mice chronically infected with T. gondii were screened. Transcriptome analysis revealed that the immune response of central nervous system of the mice with chronic infection of T. gondii was continuously activated. Seven DEGs in KYN pathway related to depression showed up-regulated transcription level.
The morphology, proliferation and stability of in vitro cultured Blastocystis were studied. Blastocystis were isolated from Blastocystis positive feces of patients with diarrhea and cultured in IMDM medium to observe the parasite morphology and proliferation. The density of Blastocystis was quantified by microscopic counting and the copy number of Blastocystis 18S small subunit ribosomal DNA (SSU rDNA) was measured by fluorescence quantitative PCR (qPCR) to analyze the proliferation of Blastocystis in the in vitro culture, and the proliferation curve was plotted to analyze the correlation between the two methods. Blastocystis were stored at 4, -20, and -80 ℃ for 1 to 7 days and 1 to 5 weeks respectively, and the Blastocystis SSU rDNA was detected by qPCR to analyze the stability. Morphological studies showed that the four common morphological forms were observed in the IMDM medium, including vacuolar form, granular form, ameboid form and cyst form; and the three reproductive modes including fission, gemmation and endodyogeny were observed. The rapid growth period of Blastocystis was observed from day 3 to 7 from the in vitro culture, and the Blastocystis density peaked on day 7. The microscopic counting and qPCR results were (2.55 ± 0.22) × 106/ml and (1.06 ± 0.10) × 106 copies/μl, respectively. Correlation analysis showed that the Pearson correlation coefficient of proliferation curve generated by microscopic counting and qPCR was 0.95, which was highly correlated. The degradation of Blastocystis began on the second day after storage at 4, -20 and -80 ℃, and the copy numbers of SSU rDNA on the seventh day were (2.75 ± 0.20) × 104, (6.84 ± 1.33) × 104 and (1.39 ± 0.06) × 105 copies/μl, respectively, which accounted for 11.65%, 28.67% and 62.42% of the copy volume (2.36 ± 0.06) × 105, (2.39 ± 0.06) × 105, (2.23 ± 0.21) × 105 copies/μl on the first day. The difference was statistically significant (F = 130.67, P < 0.05). At the fifth week, the copy numbers of SSU rDNA were (3.77 ± 0.23) × 103, (4.37 ± 0.59) × 103 and (3.86 ± 0.26) × 104 copies/μl, accounted for 2.98%, 3.41% and 28.74% of the copy volume (2.23 ± 0.21) × 105, (2.23 ± 0.21) × 105, (2.23 ± 0.21) × 105 copies/μl in the first week. The difference was statistically significant (F = 500.51, P < 0.05). The morphology of in vitro cultured Blastocystis showed diversive forms at the rapid proliferation stage. Both microscopic counting and qPCR can be used for the quantification of Blastocystis and the stability of Blastocystis stored at -80 ℃ is less than that stored at 4 and -20 ℃.
Objective To investigate the infection and molecular characteristics of Enterocytozoon bieneusi and Cyclospora cayetanensis in pet dogs and cats in Shanghai and assess the zoonotic potential. Methods Fresh fecal samples were collected from dogs and cats at a pet hospital in Shanghai from November 2021 to June 2022. Genomic DNA was extracted from the fecal samples, and nested PCR was used to amplify the internal transcribed spacer (ITS) region of the ribosomal RNA (rRNA) of E. bieneusi and the small subunit ribosomal RNA (SSU rRNA) sequence of C. cayetanensis. The positive products were subjected to bidirectional sequencing and sequence alignment by BLAST in the GenBank database. The phylogenetic tree was constructed using MEGA 11.0 software based on the neighbor-joining method. Results A total of 145 fecal samples were collected (99 from dogs and 46 from cats). The positive rates of E. bieneusi and C. cayetanensis were 4.1% (6/145) and 4.8% (7/145), respectively. The positive rates of E. bieneusi and C. cayetanensis in dogs were 3.0% (3/99) and 6.1% (6/99), respectively, while those in cats were 6.5% (3/46) and 2.2% (1/46), respectively. The ITS sequences of 6 E. bieneusi were 100% identical to human genotype A (GenBank accession number: MK982500), and the SSU rRNA sequences of 7 C. cayetanensis were 100% identical to human C. cayetanensis (GenBank accession number: KJ569533). The results of the phylogenetic tree analysis showed that all the E. bieneusi isolates belonged to the same branch as the reported human E. bieneusi genotype A isolates and fell into group 1. All the C. cayetanensis isolates belonged to the same branch as the reported human C. cayetanensis isolates. Conclusion E. bieneusi and C. cayetanensis infections were detected in pet dogs and cats in Shanghai, all of which were caused by zoonotic parasite strains.
Objective To understand the infection of Babesia in ticks parasitized on domestic animals in Xinyang, Henan Province. Methods Tick specimens were collected from domestic animals in Guangshan County and Shangcheng County, Xinyang City, Henan Province, from June to August 2022. The tick species were identified by morphology and 16S rDNA. Genomic DNA was extracted from tick specimens, and nested-PCR was performed to amplify 18S rRNA gene sequence of Babesia. The PCR products were sequenced and aligned by BLAST, and phylogenetic trees were constructed by the neighbor-joining method. Using MEGA7.0 software for homology analysis. Results A total of 335 ticks were collected, including the species of 49 Haemaphysalis longicornis, 208 Rhipicephalus microplus, 1 H. flava, 34 R. annulatus, and 43 H. punctataa, identified by morphology and PCR amplification. The PCR amplification results showed that 2 out of 335 tick samples were amplified target bands of 400 bp, and the overall positive rate was 0.6%. The BLAST comparison analysis results showed that the 18S rRNA sequence amplified from one sample had a 99.26% similarity with the sequences of B. microti from the United States (MK609547), Thailand (MG199181), and China (KU204794) in GenBank. The 18S rRNA sequence amplified from another sample shares 100% similarity with the sequences of B. gibsoni from the United States (MH620203), India (MN161136), and China (KP666166) in GenBank. The phylogenetic tree analysis revealed that the B. microti positive sample was clustered on the same branch with B. microti from Russia (KX987864), China (KU204794) and Thailand (MG199179), showing a high homology. The B. gibsoni-positive samples were clustered on the same branch with B. gibsoni from China (FJ769386), USA (MH620203) and India (KF606884), showing a close relation. Conclusion The infections of B. microti and B. gibsoni were detected in ticks parasitized on domestic animals in Xinyang area of Henan Province, which may render risk of disease transmission to humans and domestic animals.
Objective To analyze the effect of Toxoplasma gondii infection on the level of N6-methyladenosine （m6A） methylation of brain transcripts in mice. Methods C57BL/6J female mice (n = 20) were randomly divided into TgCtwh6 infection group (n = 7), LHG infection group (n = 7) and control group (n = 3 for the control of TgCtwh6 infection and LHG infection group, respectively). The TgCtwh6 infection group and the LHG infection group were inoculated by gavage with 0.2 ml brain tissue suspension (20 cysts/mouse) from the mice infected with T. gondii Chinese Ⅰ genotype wh6 strain or Chinese Ⅲ genotype LHG strain, respectively. The control group was given with the same amount of normal saline. At 15, 30 and 45 days post-infection, one mouse was randomly selected by drawing lots from each infected group, which was sacrificed under anesthesia to collect brain tissue for microscopic examination and counting of the cyst number in the cerebral cortex, hippocampus and olfactory bulb. At 45 days post-infection, the whole brain tissues of 3 mice in each group were collected, the total RNA was extracted, the genome library was prepared, the transcriptome was sequenced, differential methylation loci (DML) were screened, and differential m6A methylation sites and transcripts of mRNA in infected group and control group were recorded. The transcripts with methylation sites were analyzed by gene ontology functional annotation (GO), Kyoto Encyclopedia of Gene and Genome (KEGG), and methylated differential transcripts by gene set enrichment analysis (GSEA). Three m6A methylation modification protein genes, methyltransferase like 3 (METTL3), fat mass and obesity-associated protein (FTO) and YTH domain family 3 (YTHDF3), were selected, and their relative transcription levels were detected by real-time fluorescence quantitative reverse transcription PCR (qRT-PCR) using the glyceraldehyde-3-phosphate dehydrogenase gene as an internal reference. Results All the mice in the infection group were infected successfully. On day 45 after infection, the cysts in cerebral cortex, hippocampus and olfactory bulb were (5 676 ± 10), (4 773 ± 9) and (243 ± 10), respectively. A total of 760 650 methylation sites were detected in the infection group and control group. Among them, the methylation levels of four m6A motifs, GGACA, GGACC and ACGAT, accounted for 16.8% (127 923/760 650), 4.6% (35 164/760 650), 3.8% (28 983/760 650) and 0.4% (3 122/760 650), respectively. A total of 127 016 transcripts with high methylation sites were detected, including 71 727, 27 754, 24 556 and 2 979 transcripts with GGACA, GGACC, GGACT and ACGAT, respectively. The total number of DML sites for four m6A motifs, including ACGAT, GGACA, GGACC and GGACT, in the transcriptome of the infected and control group mice brain tissue was 9 233, including 4 832 high methylation sites and 4 851 low methylation sites. The GO analysis showed that the transcripts where DML was located were significantly enriched in cellular processes of biological processes, cell of cellular components, and binding in molecular functions. The KEGG enrichment analysis of the transcripts where DML was located showed that the intermolecular interaction network was mainly enriched in signaling pathway such as lysosome, spliceosomes, dopamine synapses. The GO enrichment analysis of biological processes showed that in the transcripts where DML was located, some transcripts related to biological processes were mainly enriched in processes such as protein glycosylation, negative regulation of neuronal apoptosis and proteion import into nucleus. The GSEA analysis showed the differential methylation between the control group and the two infection groups was highly enriched in transcriptome subsets related to the negative regulation pathway of fibroblast proliferation and Hippo signaling pathway. Among them, four key transcripts, ENSMUST00000105393, ENSMUST00000006523, ENSMUST00000055261 and ENSMUST00000038658, were screened, which encode costimulatory factor ligand (ICOSL), cysteine-rich intestinal protein 1 (CRIP1), MOB kinase activator 1A201 (MOB1A201) and MOB1A202, respectively. Compared with the control group, the expression of these four genes in TgCtwh6 infection group and LHG infection group were all up-regulated. The results of qRT-PCR showed that the relative expression level of mettl3 mRNA in brain tissue of TgCtwh6 infection group and LHG infection group was 5.47 ± 1.09, 1.63 ± 0.06, respectively, which was significantly different from that of the control group (1.01 ± 0.11) (t = 4.05, 5.03; both P < 0.05). The relative expression level of ythdf3 mRNA in the control group was 3.57 ± 0.08 and 1.80 ± 0.25, respectively, which was significantly different from that in the control group (1.01 ± 0.11) (t = 18.95, 2.85; both P < 0.05). The relative expression level of fto mRNA was 0.41 ± 0.04, 0.60 ± 0.12, respectively, which was significantly different from the control group (1.00 ± 0.06) (t = 7.67, 2.99; both P < 0.05). The qRT-PCR results were consistent with the transcriptional trend obtained by transcriptional sequencing. Conclusion Chronic T. gondii infection may enhance the m6A methylation in mouse brain transcripts, leading to changes in the biological processes and signal pathways related to host mental behavioural through modifying and regulating key differential transcripts including icosl, crip1 and mob1a.
Zoonotic diseases pose a serious threat to human health and ecological security. Climate change facilitates the crossover and spread of zoonotic diseases by affecting pathogens, hosts, vectors, and human activities, therefore threatening global public health. This article summarizes the impact of climate change on the spread of zoonotic diseases and explores the effective strategies based on the One Health approach to protect human health and safety.
To understand the work progress in nationwide control of echinococcosis, we summarize the experience and find the existing problems through descriptive analysis on the national control data in 2022. As of the end of 2022, there were 370 echinococcosis endemic counties (city, district, banner) covering 29 926 villages in China, having a total of 25 227 echinococcosis cases with an average prevalence of 58.35 per 100 000 （25 227/43 232 609), among them, 15 554 cases were of cystic echinococcosis, 8 169 cases alveolar echinococcosis, 255 mixed infection, and 1 249 cases pending; 1 270 cases were newly found, including 991 cases of cystic echinococcosis, 89 alveolar echinococcosis, 5 mixed infection, and 185 pending; revealing 102 cases were at age < 12, and 1 168 case were of age ≥ 12. In 2022, population screening by abdominal ultrasound scanning was performed in all endemic provinces (autonomous regions) for 3 576 121 person/times. Of them, 751 440 person/examinations were performed for the residents younger than 12, and 2 824 681 person/times were performed for residents older than 12. The serological examination was carried out for the suspected individuals for 17 404 person/times. According to data from 370 surveillance sites in 2022, the prevalence in residents younger than 12 was 0.02% (60/287 437) by ultrasound screening, with 40.00% (24/60) newly diagnosed cases. The prevalence in the residents older than 12 was 0.29% (772/270 407) in type Ⅰ and type Ⅱ endemic counties (city, district, banner). The newly diagnosed cases accounted for 8.29% (64/772) of the detected patients. In 2022, 18 354 patients received drug treatment, of which 16 625 patients received liver and kidney function tests or management against adverse reactions. A total of 1 418 patients received surgical treatment, including 69.82% (990/1 418) for cystic echinococcosis, and 26.52% (376/1 418) for alveolar echinococcosis. In 2022, the follow-up results showed that 999 cases were cured, 20 599 cases responded to the treatment, 2 546 cases failed in the treatment, 374 cases died (the causes of the deaths were not echinococcosis), 239 cases were excluded, 372 cases were lost in follow-up, 884 cases had not completed the follow-up, and 170 cases migrated to other places. In 2022, there were 2 478 608 dogs in the endemic townships (towns) nationwide, of which 2 250 694 were registered for management. Deworming work was conducted for dogs in 34 646 villages, with 24 289 457 deworming times. Wild canines were demormed by delivering 119 473 drug doses. A total of 394 851 fecal samples from domestic dogs were collected and detecteded, of which 1 756 were found positive for Echinococcus coproantigen, with a positive rate of 0.44% (1 756/394 851). Of the wild canines, 62 884 field fecal samples were collected and detected, among which 1 201 were found positive for Echinococcus coproantigen, with a positive rate of 1.91% (1 201/62 884). A total of 117 303 slaughtered livestock were randomly examined, among which 1 038 were diseased, with a prevalence of 0.88%. A total of 43 705 field rodents were examined, among which 403 were diseased, with a prevalence of 0.92%. Although the endemic of echinococcosis in China has been essentially controlled, there remain many difficulties and challenges in control work. It is imperative to continuously strengthen the prevention and control measures for echinococcosis, raise the disease prevention awareness, explore optimizing the control strategy, exert the role of regional joint prevention and control mechanisms and fully implement comprehensive measures to further control the prevalence of echinococcosis.
Objective To understand the epidemic situation of visceral leishmaniasis in China in 2022 and provide scientific basis for formulating prevention and control strategy. Methods Data of visceral leishmaniasis in 2022 was collected from the web-based National Infectious Diseases Reporting Information Management System. After excluding suspected cases, duplicates and cutaneous leishmaniasis cases, a visceral leishmaniasis database was established and underwent descriptive epidemiological analysis with Microsoft Excel 2016. Results A total of 239 cases of visceral leishmaniasis were reported in 104 counties of 11 provinces (autonomous regions and municipalities) in 2022, among them 191 cases were reported from mountain-type zoonotic visceral leishmaniasis endemic areas, 4 cases from desert-type zoonotic visceral leishmaniasis endemic areas, 2 cases from anthroponotic visceral leishmaniasis endemic areas and 42 cases were imported from non-endemic areas. These cases were mainly distributed in Shanxi (110 cases), Shaanxi (34 cases), Henan (23 cases) and Hebei (23 cases), the total reported accounting for 79.30% (190/239) of the overall total in China. A total of 197 local transmitted cases were reported from 42 endemic counties and other 63 counties were of non-endemic areas, reporting 39 imported cases. Pingding County, outer suburbs of Yangquan City, Jingxing County and Huazhou District were the major endemic counties with 25, 15, 16 and 10 cases reported respectively, the total accounting for 27.62% (66/239) of the overall total cases in China. Recurrence endemic counties of visceral leishmaniasis were mainly concentrated in Shanxi (Qinshui County, Gaoping City, Fushan County, Houma City, Yicheng County, Jiangxian County, Yuanqu County, Wenshui County), Hebei (Mining district of Jingxing County, Zanhuang County, Lincheng County, Xindou District), Henan (Yanshi District, Shangjie District, Qibin District), Beijing (Changping District) and Xinjiang (2th regiment farm) in 2022, with a total of 23 local cases reported. The peak incidence occurred in July. The ratio of male to female cases was 1 ∶ 0.38. Farmers, infants and young children are the high-risk population of visceral leishmaniasis, accounted for 53.97% (129/239) and 16.74% (40/239) of the total cases respectively. The reported cases at age of ≥ 15 accounted for 81.17% (194/239). Conclusion Visceral leishmaniasis is at a low prevalence status in China, whereas the endemic area is gradually expanding, and the number of cases is gradually increasing in mountain-type zoonotic visceral leishmaniasis endemic area.
Objective To understand the distribution pattern of the neurotransmitter 5-hydroxytryptamine (5-HT) in Clonorchis sinensis. Methods The metacercariae of C. sinensis were isolated from infected Pseudorasbora parva fish by digestion, and used to infect Kunming mice (40-50 metacercariae/mouse) by gavage. Mice were euthanized 10, 20 and 30 days after infection, and the parasites were collected from the liver portal vein. The development status of worm organs at different stages were observed after staining with acetic carmine under optical microscope, at the same time, 5-HT immunofluorescence staining was performed to observe the distribution of 5-HT using laser scanning confocal fuorescence microscope. Results Under the optical microscope, the acetic acid magenta staining showed that in the C. sinensis d10 larva, the digestive and excretory organs were basically mature, the reproductive organs were not fully mature, and scattered eggs could be seen in the uterus. In the d20 larva, the reproductive organs in the worm were matured, the number of eggs in the uterus increased, and the testis branches increased. In the d30 adult worm, the digestive, excretory and reproductive systems were all mature, the eggs were more closely arranged in the uterus than the larvae, and the testis were enlarged and branched obviously. Under laser scanning confocal microscopy, the 5-HT immunofluorescence staining showed that in the d10 larva, the fluorescence staining was strong in the central parts of the body through the system segment, nerve union and oral sucker and was distributed in a dotted pattern within the internal organs in the body. In the d20 larva, a small number of nerve cells were observed in the nerve union near the ventricle sucker. Besides the central nervous system, 5-HT fluorescence staining appeared in the excretory sac, excretory pore and reproductive organs of the body. In the d30 adult worm, 5-HT was found in the digestive, excretory and reproductive organs of the worm, and the fluorescence staining of the testis was stronger than that of intestinal branches and oral sucker, and there were more nerve cells in the worm. Conclusion 5-HT is widely distributed in the body of C. sinensis, mainly in the organs and viscera rich in muscle tissue. There were differences in the distribution of 5-HT in different organs at the same developmental stage, and differences were also seen in the same type of organs at different developmental stages.
A 53-year-old male patient, who is a farmer, lived in Tanchang of Gansu, was treated at the Neurology Clinic of Gansu Provincial People’s Hospital due to “headache, dizziness, nausea, and vomiting for one week” on November 19, 2021. On admission, CT plain scan of the brain showed nodular and slightly high-density lesions in the transparent septum, while MRI of the head showed mild ventricular dilation. Lumbar puncture result showed that the intracranial pressure was 180 mmH2O (1 mmH2O = 9.779 Pa). The cerebrospinal fluid laboratory examination showed that the total protein in cerebrospinal fluid was 0.74 g/L. The second lumbar puncture result showed that the intracranial pressure was 300 mmH2O and the patient was transferred to neurosurgery on the following day. The patient was drowsy with high cranial pressure and unable to speak correctly upon awakening. Repeat CT scan showed lateral ventricular dilatation, which suggested high risks of hydrocephalus and cerebral hernia. The patient had eaten undercooked meat in recent years and a history of liver echinococcosis. To reduce patient’s cranial pressure, ventriculocentesis was performed on November 26 and the patient got the intensive care after surgery. The serum samples were positive for cysticercosis IgG and Toxoplasma gondii IgG. Therefore, the patient was treated with praziquantel (400 mg/8 h) and albendazole (0.4 g/d) for 3 courses (7 d/course, each treatment interval of 5 d). Endoscopic third ventriculostomy (ETV) was performed for the hydrocephalus on December 13. The patient got the antiparasitic therapy after ETV and lumbar ampullary drainage was performed at the same time, but the condition was not improved. The ventriculocentesis was performed twice on December 28 and January 11, 2022 respectively to reduce patient’s cranial pressure. The patient was treated with antiparasitic therapy and tigecycline (50 mg/12 h) and sulperazon (3 g/8 h). The patient’s intracranial infection indicators turned negative on January 25 and enhanced brain MRI showed no obvious tapeworm lesions. Laparoscopic assisted ventriculoperitoneal shunt was performed on January 27. The patient’s was consciouse and the cranial CT scan showed improved hydrocephalus after the surgery. The patient was discharged from the hospital with clear consciousness and no obvious dizziness, headache, nausea, vomiting or seizures on February 11. The patient recovered well after 3 months.
Objective To analyze the epidemiological characteristics of malaria in Jiangxi Province from 2015 to 2022, to provide scientific basis for formulating prevention and control strategy. Methods Malaria cases and statistical data were collected from the National Infectious Diseases Reporting and Information Management System and the National Information Management System for Parasitic Diseases Control and Prevention, for those cases locally reported and confirmed during January 1st, 2015 to December 31th, 2022 in Jiangxi Province. The collected malaria cases were classified as confirmed cases (microscopic examination and/or PCR test positives confirmed by provincial reference laboratory for malaria diagnosis), clinically diagnosed cases (blood examination positive found by medical institutions, but microscopic examination and PCR test by provincial reference laboratory were negative), and severe case (with complications). Statistical analysis was performed by LSD-t and mann-whitney Test using SPSS 26.0 software. Results From 2015 to 2022, a total of 241 malaria cases were reported in Jiangxi Province, all of which were imported malaria and no deaths were reported. The reported cases in each year were 53, 52, 30, 41, 46, 12, 3, and 4, respectively. Among them, there were 240 confirmed cases and 1 clinically diagnosed case (reported in 2015). Among the confirmed cases, there were 153 cases of falciparum malaria (63.75%), 50 cases of vivax malaria (20.83%), 31 cases of ovale malaria (12.91%), 5 cases of malariae malaria (2.08%), and 1 case of mixed infection of falciparum malaria and vivax malaria (0.41%). The sources of infection are distributed in 34 countries including Africa, Asia, and Oceania, with Africa (a total of 27 countries) accounting for 92.53% (223/241). There are case reports in each month, with a cumulative increase in cases in January, June, and September, and a decrease in cases in October, November, and December. From 2015 to 2019 (before the COVID-19 epidemic) and from 2020 to 2022 (during the COVID-19), the monthly average number of reported cases was 3.7 and 0.53 respectively (t = 6.369, P < 0.05). All 11 prefectures in Jiangxi Province have reported cases, with the top 3 prefectures reporting cases being Nanchang City (51.45%, 124/241), Ganzhou City (15.77%, 38/241), and Yichun City (7.05%, 17/241); The top three counties (cities, districts) reporting cases are Qingshanhu District of Nanchang City (33.19%, 80/241), Donghu District of Nanchang City (9.54%, 23/241), and Zhanggong District of Ganzhou City (7.88%, 19/241). Among the reported cases of malaria, males accounted for 95.44% (230/241) and females accounted for 4.56% (11/241). The age of onset was mainly concentrated between 20 and 50 years old (97.11%, 234/241), mainly reported by medical institutions and disease control centers at or above the county level (99.59%, 240/241). The median interval between onset and initial visit is 1 day; The average number before the COVID-19 was (2.76 ± 5.00) d, and the average number during the epidemic was (1.79 ± 1.81) d, with no statistically significant difference (Z = -0.155, P > 0.05). The median time interval between initial diagnosis and diagnosis is 2 days; The average number before the COVID-19 was (3.36 ± 3.30) d, and the average number during the epidemic was (2.74 ± 2.90) days, with no statistically significant difference (Z = -0.103, P > 0.05). There were 28 severe cases (11.62%) and 213 non severe cases (88.38%). There was no statistically significant difference in the interval between onset and diagnosis (median 6 d) between severe cases and non severe cases (median 4 d) (Z = -1.242, P > 0.05). Conclusion In the malaria post-elimination stage, there remains retransmission risk caused by imported cases, thus it is imperative to continuously strengthen surveillance to prevent from the occurance of severe cases and deaths.
The data of reported echinococcosis cases in Jiangsu Province from 2015 to 2022 were collected from the Chinese Disease Prevention and Control Information System and epidemiological investigations were conducted on individual cases, while infection of intermediate and terminal hosts was monitored. Two key areas of Liyang City and Yixing City, where suspected local cases have occurred in the past were selected to conduct an investigation on the infection status of intermediate and final hosts of echinococcosis. Every year, 100-200 sheep organ samples were collected from slaughterhouses in both cities, and visceral dissection was used to check the infection status of Echinococcus. Fecal samples from at least 100 dogs in 1-3 administrative villages were collected, and canine fecal Echinococcus antigen was detected by ELISA. A descriptive analysis was performed using PASW 18.0 to analyze the time, region, population distribution, treatment status, and surveillance results of intermediate and terminal hosts. From 2015 to 2022, a total of 29 cases of echinococcosis were reported in Jiangsu Province, of which 62.07% (18/29) were confirmed cases, and 37.93% (11/29) were clinically diagnosed cases. There were 14 suspected local cases and 15 imported cases. Twelve of the imported cases were from Xinjiang, China, 2 cases from Xizang, China and 1 case from Pakistan. Thirteen reported cases, which was the highest in number, were from Changzhou (10 of which are suspected local cases in Liyang). Among the 29 reported cases, 34.48% (10/29) were male and 65.52% (19/29) were female, 51.72% (15/29) were 40-60 years old, and 41.38% (12/29) were farmers. All cases were cystic echinococcosis, and the symptoms of upper abdominal discomfort accounted for 51.72% (15/29), and the main parasitic site was the liver (93.10%, 27/29). 75.86% (22/29) of the cases received surgical treatment. The fecal antigen-positive rate of canine Echinococcus was 0.74% (15/2 028), and suspected Echinococcus eggs were found in two canine feces. From 2015 to 2022, echinococcosis cases were reported annually in Jiangsu Province, and in addition to imported cases, there were also suspected local cases. The transmission chain of echinococcosis may exist in local areas. Therefore, follow-up surveillance of echinococcosis should be strengthened, and screening and health education should be actively carried out in key areas and populations.
Objective To explore the diagnosis and differentiation between Opisthorchis viverrin and Opisthorchis sinensis infection through morphological examination of small trematode eggs, egg internal transcribed space 2 (ITS-2) sequence amplification and epidemiological investigation. Methods The epidemiological data of 6 trematode egg positive cases diagnosed by the modified Kato-Katz thick smear method were collected, and positive fecal samples were collected for 3 consecutive days for collection of eggs by washing with water and sedimentation prior to microscopy. The eggs DNA was extracted, amplified and sequenced using universal primers for the rDNA ITS-2 region. The product sequences were aligned in the NCBI database to identify the species, and a phylogenetic tree based on ITS-2 sequences was constructed using the neighbor-joining method. The diagnosis was reached based on data analysis of the egg morphology, gene sequence alignment and epidemiological findings. Results Microscopically, the eggs in the fecal samples of cases 1 to 3 were similar to those of cases 4 to 6 in the shell, morphology, color, egg cover, acromion, hair and small wart at the bottom of the eggs. The egg length, width and aspect ratio in the fecal samples from cases 1 to 3 were (26.94 ± 2.28) μm, (14.43 ± 1.22) μm, 1.88 ± 0.18. Compared with (29.70 ± 1.21) μm, (14.36 ± 0.70) μm and 2.07 ± 0.14 in cases 4 to 6. The difference between length and aspect ratio was significant (t = 4.318, 4.816; P < 0.01). The former is shorter and wider than the latter, The Forde-Meleni index (FMI) of the two eggs was 5 676.69 ± 1 317.97 and 6 134.25 ± 626.27 (t = 0.428, P > 0.05). In cases 1 to 3, PCR and sequencing results showed that the ITS-2 fragments were 355, 362 and 359 bp, respectively, with the sequence identity of 99.14%, 98.86% and 99.39% to C. sinensis (GenBank No. MK886663). In cases 4 to 6, ITS-2 gene fragments were 394, 395 and 396 bp, respectively, with the sequence identity of 100%, 99.49% and 100% to O. viverrin (GenBank No. OK103575.1). The phylogenetic tree showed that the ITS-2 sequence of the eggs in cases 1 to 3 clustered on a branch with C. muscidiode and the ITS-2 sequence of the eggs in cases 4 to 6 clustered on a branch with C. sinensis. The epidemiological investigation results showed that cases 1 to 3 were all foreign migrant women from Cambodia who had the habit of eating pickled freshwater fish since childhood; cases 4 to 6 were all local farmers. Combining the morphological, molecular biology and epidemiological findings, cases 1 to 3 were diagnosed as imported testicular fluke infection, and cases 4 to 6 were C. sinensis infection. Conclusion Combining morphological detection as well as epidemiological investigation and analysis, trematode egg ITS-2 amplification and sequencing could effectively differentiate C. sinensis and O. viverrin in infection.
Objective To investigate the endemic status of Fasciola spp. infection in cattle and sheep in Wushen Banner of Inner Mongolia. Methods From August 2022 to March 2023, blood and fecal samples from cattle and sheep were collected from Galutu, Sulide Sumu, Wulantaolegai, Wushenzhao and Tuke towns in Wushen Banner to detect serum Fasciola hepatica antibodies by using indirect ELISA. Cattle and sheep feces samples corresponding to some serum antibody detection samples were randomly selected, in which the parasite eggs were qualitatively examined by egg sedimentation method. The consistency test was performed for the findings from serum and fecal examination. In April 2023, Fasciola spp. adult worms and eggs were collected from the carcasses livers and bile of cattle and sheep that died of Fasciola spp. infection in Galutu, Sulidesumu and Tuke towns. The DNA of the worms and eggs was extracted to amplify ribosomal transcriptional spacer sequence 2 (ITS2) by PCR and sequenced. Sequence alignment was then performed by BLAST with NCBI database. The phylogenetic tree was constructed by the neighbour-joining method. SPSS 25.0 software was used for statistical analysis, and the chi-square test was used to compare the positive rates in different regions. Results A total of 825 serum samples were collected from cattle and sheep, in which 295 samples were positive for serological antibodies, with a total sero-positive rate of 35.8% (295/835), of which the sero-positive rates of sheep and cattle were 34.7% (227/655) and 40.0% (68/170), respectively. The sero-positive rate of sheep was the highest at 81.8% (45/55) in Sulide Sumu, and the lowest at 22.0% (27/123) in Wushenzhao. There was a significant difference in the sero-positive rate of antibodies in different regions (χ2 = 73.93, P < 0.05). The sero-positive in bovine was the highest at 65.1% (28/43) in Garutu and lowest at 19.2% (5/26) in Tuke. There was a significant difference in the sero-positive rate of antibodies in different regions (χ2 = 21.50, P < 0.05). A total of 88 fecal samples (59 sheep fecal samples and 29 bovine fecal samples) were detected, among which 1 sheep and 5 bovine fecal samples were positive for F. hepatica eggs, and 4 bovine fecal samples were positive for Paramphistomum eggs. Among the 88 samples, the positive rate of anti-F. hepatica antibody was 29.5% (26/88), and the positive rate of F. hepatica eggs was 6.8% (6/88). The kappa value was 0.086, indicating weak consistency between the two methods. Adult worms and eggs were collected from 2 dead cattle and adult worms from 2 dead sheep. The DNA were extracted from 5 adult worms and 2 eggs samples. A total of 7 bands of approximately 550 bp were obtained by PCR amplification for sequencing. BLAST alignment analysis showed that the identities of 4 obtained sequences were 92.97%, 96.62%, 95.74% and 95.91% with the ITS2 sequences (GenBank: HQ700438) of F. gigantica from China, respectively, and identified as F. gigantica. The identities of the other three obtained sequences were 95.69%, 99.80% and 99.23% with the ITS2 sequences (GenBank: JF496717) of F. hepatica from China, respectively, and identified as F. hepatica. The phylogenetic tree showed that the F. hepatica identified in this study clustered on the same branch with the F. hepatica from China (GenBank: JF496717), Kenya (GenBank: MZ396926) and Egypt (GenBank: MW620063). The F. gigantica identified in this study clustered in the same branch with the F. gigantica from China (GenBank: HQ700438) and India （GenBank: OL691113）. Conclusion F. hepatica and F. gigantica infections are present in both cattle and sheep in Wushenqi area of Inner Mongolia, and the infection rate of Fasciola spp. was relatively high.
Objective To probe the immunomodulatory effects of Cryptosporidium parvum micronemal glycoprotein 900 (GP900) 829-1 099aa fragments (GP900829-1099) on mouse macrophages (RAW264.7 cells) through the nuclear factor kappa-B (NF-κB)/mitogen-activated protein kinase (MAPK) signalling pathways. Methods The amino acid sequence of GP900829-1099 was obtained from the NCBI database for bioinformatics analysis. Sequence fregment of gp900829-1099 was amplified with PCR using cDNA from C. parvum oocysts as a template. After gel extraction, gp900829-1099 were inserted into the pET-32a-gp900829-1099 vector and transformed into BL21 competent cells for induced expression. The recombinant proteins were purified and filtered, concentrated by ultrafiltration and further cleaned by removing endotoxin. The GP900829-1099 recombinant protein at concentration of 0.16, 0.80, 4.00, 20.00, 100.00 and 500.00 μg/ml was applied to stimulate RAW264.7 for 24 h, subsequently, its regulatory effect on the cell viability was detected by CCK-8 assay. Using PBS as the control group and lipopolysaccharide（1 μg/ml）as the positive control group, the recombinant protein at the concentration of 0.16, 0.80 and 4.00 μg/ml was applied to stimulate the RAW264.7 cells for 24 h to detect CD86 expression by flow cytometry. qPCR was used to detect the relative transcription levels of IL-6 and TNF-ɑ mRNA in RAW264.7 cells. ELISA was used to detect the expression levels of IL-6 and TNF-ɑ in the supernatant of RAW264.7 cell culture. The phosphorylation levels of P65 protein and extracellular regulated protein kinase (ERK) in NF-κB and MAPK signaling pathways were detected by Western blotting assay. Results In the secondary structure of GP900829-1070 proteins β corners account for 9.23%, irregular curls account for 64.58%, and contain abundant T and B cell antigenic epitopes. The molecular mass of the expressed recombinant protein GP900829-1099 was consistent with the theoretical value. The CCK8 results showed no toxic effect on the cells, and the subsequent working concentrations were selected as 0.16, 0.80 and 4.00 μg/ml. Flow cytometry results showed that the positive expression rate of CD86+ was (13.500 ± 0.815)%, (18.670 ± 0.657)% and (20.470 ± 1.271)%, respectively, and the latter two were higher than that of the blank control group (14.500 ± 0.872)% (t = 3.818, 3.872; both P < 0.05). qPCR results showed that the relative transcription levels of IL-6 mRNA in macrophages in 0.16, 0.80 and 4.00 μg/ml groups were 1.409 ± 0.050, 2.052 ± 0.098 and 3.284 ± 0.097, respectively, which were higher than those in the blank control group (1.010 ± 0.097) (t = 3.700, 7.595, 16.700; all P < 0.05). The TNF-ɑ mRNA relative transcription levels in macrophages of the three concentration gradient groups were 1.077 ± 0.034, 1.440 ± 0.021 and 2.378 ± 0.037, respectively, with the latter two being higher than that of the blank control group (1.000 ± 0.025) (t = 13.380, 30.850; both P < 0.01). ELISA results showed that the expression levels of IL-6 cytokines in macrophage culture supernatants were (535.400 ± 17.230), (572.800 ± 8.286) and (555.600 ± 23.940) mg/L in the 0.16, 0.80 and 4.00 μg/ml groups, respectively, which was higher than that in the blank control group [(454.400 ± 18.630) mg/L] (t = 3.193, 5.809, 3.339; all P < 0.05); the expression levels of TNF-ɑ cytokine were (351.800 ± 12.270), (386.400 ± 10.250) and (489.800 ± 10.540) mg/L, respectively, and the latter two were higher than that of the blank control group [(324.200 ± 11.070) mg/L] (t = 4.125, 10.830; both P < 0.01). Western blotting results showed that the relative expression levels of p-P65 protein and p-ERK protein in macrophages in 0.16, 0.80 and 4.00 μg/ml groups were 2.294 ± 0.254, 1.714 ± 0.205, 1.877 ± 0.309 and 1.522 ± 0.054, 1.760 ± 0.066, 1.582 ± 0.027, all were higher than that of the blank control group (1.0 ± 0.0) (t = 5.100, 3.489, 2.836 and 9.737, 11.450, 21.900; all P < 0.05). Conclusion GP900829-1070 recombinant protein stimulated macrophages at different concentrations, which activates NF-κB and MAPK signalling pathways to induce activation of macroopages and promote the transcription of TNF-α and IL-6 mRNA, thereby, participating in macrophage immunomodulation.
