A point-of-care testing assay for clonorchiasis using a EuNPs-CsTR1 fluorescent probe-based immunoassay.

Clonorchis sinensis is one of the most important fish-borne zoonotic parasitic worms in humans, and is distributed in several countries with more than 15 million people infected globally. However, the lack of a point-of-care testing (POCT) method is still the critical barrier to effectively prevent clonorchiasis. With the application of novel fluorescent nanomaterials, the development of on-site testing methods with high signal enhancement can provide a simple, precise and inexpensive tool for disease detection. In this study, Eu-(III) nanoparticles (EuNPs) were used as indicative probes, combined with C. sinensis tandem repeat sequence 1 (CSTR1) antigen to capture specific antibodies. Afterward, the complex binds to mouse anti-human IgG immobilized on the test line (T-line) producing a fluorescent signal under UV light. The EuNPs-fluorescent immunoassay (EuNPs-FIA) was successfully constructed, allowing sample detection within 10 min. It enabled both qualitative determination with the naked eye under UV light and quantitative detection by scanning the fluorescence intensity on the test line and control line (C-line). A total of 133 clinical human sera (74 negative, 59 clonorchiasis, confirmed by conventional Kato-Katz (KK) methods and PCR via testing fecal samples corresponding to each serum sample) were used in this study. For qualitative analysis, the cut-off value of fluorescence for positive serum was 31.57 by testing 74 known negative human samples. The assay had no cross-reaction with other 9 parasite-infected sera, and could recognize the mixed infection sera of C. sinensis and other parasites. The sensitivity and specificity of EuNPs-FIA were both 100% compared with KK smear method. Taking advantage of its high precision and user-friendly procedure, the established EuNPs-FIA provides a powerful tool for the diagnosis and epidemiological survey of clonorchiasis.

Clonorchiasis and opisthorchiasis: epidemiology, transmission, clinical features, morbidity, diagnosis, treatment, and control.

Clonorchis sinensis, Opisthorchis viverrini, and Opisthorchis felineus are important liver flukes that cause a considerable public health burden in eastern Asia, southeastern Asia, and eastern Europe, respectively. The life cycles are complex, involving humans, animal reservoirs, and two kinds of intermediate hosts. An interplay of biological, cultural, ecological, economic, and social factors drives transmission. Chronic infections are associated with liver and biliary complications, most importantly cholangiocarcinoma. With regard to diagnosis, stool microscopy is widely used in epidemiologic surveys and for individual diagnosis. Immunologic techniques are employed for screening purposes, and molecular techniques facilitate species differentiation in reference laboratories. The mainstay of control is preventive chemotherapy with praziquantel, usually combined with behavioral change through information, education and communication, and environmental control. Tribendimidine, a drug registered in the People's Republic of China for soil-transmitted helminth infections, shows potential against both C. sinensis and O. viverrini and, hence, warrants further clinical development. Novel control approaches include fish vaccine and biological control. Considerable advances have been made using multi-omics which may trigger the development of new interventions. Pressing research needs include mapping the current distribution, disentangling the transmission, accurately estimating the disease burden, and developing new diagnostic and treatment tools, which would aid to optimize control and elimination measures.

A rapid and visual detection assay for Clonorchis sinensis based on recombinase polymerase amplification and lateral flow dipstick.

BACKGROUND: Fish-borne zoonotic clonorchiasis, caused by Clonorchis sinensis, is an emerging public health problem in several countries with more than 15 million people infected globally. However, a lack of accurate point-of-care (POC) diagnostic tests in resource-limited areas is still a critical barrier to effective treatment and control of clonorchiasis. The development of the recombinase polymerase amplification(RPA) assay, a POC diagnostic test based on the amplification of pathogen DNA, has provided a new, simple and inexpensive tool for disease detection with high sensitivity and specificity. METHODS: A novel RPA method was developed based on specific primers and probes, and combined with the dipstick, to allow for the rapid and intuitive detection of C. sinensis through the amplification of the mitochondrial cytochrome c oxidase subunit 1 (COX1) gene. The lower limit of detection for the combined RPA/lateral flow dipstick (RPA-LFD) assay was evaluated using dilutions of the target DNA sequence. Cross-reactivity was evaluated using genomic DNA from 10 additional control parasites. Forty human clinical stool samples were tested to verify its performance. RESULTS: The evaluated primers designed from the C. sinensis COX1 region can be used to detect adult worms, metacercariae, and eggs at 39 °C within 20 min, and the results can be visually observed using the LFD. The detection limit of pathogen genomic DNA was as low as 10 fg, and the number of metacercaria(e) in fish and egg(s) in faeces were both as low as one. This improved the sensitivity of low-infection detection tremendously. The test is species-specific, and no other related control parasites were detected. In human stool samples with eggs per gram (EPG) > 50, the RPA-LFD assay was performed consistent with conventional Kato-Katz (KK) and PCR methods. CONCLUSION: The established RPA-LFD assay provides a powerful tool for the diagnosis and epidemiological survey of C. sinensis from human and animal samples, and has important implications for the effective control of clonorchiasis.

Genome-wide exploration reveals distinctive northern and southern variants of Clonorchis sinensis in the Far East.

Clonorchis sinensis is a carcinogenic liver fluke that causes clonorchiasis-a neglected tropical disease (NTD) affecting ~35 million people worldwide. No vaccine is available, and chemotherapy relies on one anthelmintic, praziquantel. This parasite has a complex life history and is known to infect a range of species of intermediate (freshwater snails and fish) and definitive (piscivorous) hosts. Despite this biological complexity and the impact of this biocarcinogenic pathogen, there has been no previous study of molecular variation in this parasite on a genome-wide scale. Here, we conducted the first extensive nuclear genomic exploration of C. sinensis individuals (n = 152) representing five distinct populations from mainland China, and one from Far East Russia, and revealed marked genetic variation within this species between "northern" and "southern" geographical regions. The discovery of this variation indicates the existence of biologically distinct variants within C. sinensis, which may have distinct epidemiology, pathogenicity and/or chemotherapic responsiveness. The detection of high heterozygosity within C. sinensis specimens suggests that this parasite has developed mechanisms to readily adapt to changing environments and/or host species during its life history/evolution. From an applied perspective, the identification of invariable genes could assist in finding new intervention targets in this parasite, given the major clinical relevance of clonorchiasis. From a technical perspective, the genomic-informatic workflow established herein will be readily applicable to a wide range of other parasites that cause NTDs.

Structure-based virtual screening and molecular dynamics of potential inhibitors targeting sodium-bile acid co-transporter of carcinogenic liver fluke Clonorchis sinensis.

BACKGROUND: Clonorchis sinensis requires bile acid transporters as this fluke inhabits bile juice-filled biliary ducts, which provide an extreme environment. Clonorchis sinensis sodium-bile acid co-transporter (CsSBAT) is indispensable for the fluke's survival in the final host, as it circulates taurocholate and prevents bile toxicity in the fluke; hence, it is recognized as a useful drug target. METHODOLOGY AND PRINCIPAL FINDINGS: In the present study, using structure-based virtual screening approach, we presented inhibitor candidates targeting a bile acid-binding pocket of CsSBAT. CsSBAT models were built using tertiary structure modeling based on a bile acid transporter template (PDB ID: 3zuy and 4n7x) and were applied into AutoDock Vina for competitive docking simulation. First, potential compounds were identified from PubChem (holding more than 100,000 compounds) by applying three criteria: i) interacting more favorably with CsSBAT than with a human homolog, ii) intimate interaction to the inward- and outward-facing conformational states, iii) binding with CsSBAT preferably to natural bile acids. Second, two compounds were identified following the Lipinski's rule of five. Third, other two compounds of molecular weight higher than 500 Da (Mr > 500 Da) were presumed to efficiently block the transporter via a feasible rational screening strategy. Of these candidates, compound 9806452 exhibited the least hepatotoxicity that may enhance drug-likeness properties. CONCLUSIONS: It is proposed that compound 9806452 act as a potential inhibitor toward CsSBAT and further studies are warranted for drug development process against clonorchiasis.

[Clinical characteristics of biliary obstruction patients with Clonorchis sinensis infections].

OBJECTIVE: To analyze the clinical characteristics of biliary obstruction patients with Clonorchis sinensis infections, so as to provide insights into the clinical diagnosis and therapy of this comorbidity. METHODS: A total of 45 biliary obstruction patients with C. sinensis infections that were admitted to the Second Affiliated Hospital of Harbin Medical University from October 2012 to September 2019 were enrolled, and 45 patients with biliary obstruction alone at the hospital during the same study period were recruited as controls. Univariate analysis was performed to analyze the epidemiological characteristics, clinical manifestations, laboratory examination results and imaging manifestations related to C. sinensis infection, and the statistically significant univariate was used as an independent variable for multivariate logistic regression analysis. RESULTS: Compared with biliary obstruction alone, males (91.11% vs. 46.67%; χ2 = 20.737, P < 0.01) and rural areas (62.22% vs. 22.22%; χ2 = 14.757, P < 0.01) showed a significantly higher proportion in biliary obstruction patients with C. sinensis infections. The major clinical symptoms involved in had jaundice (45 cases, 100%) and abdominal pain (40 cases, 88.89%) in C. sinensis-infected patients. Logistic regression analysis showed that males [odds ratio (OR) = 10.717, 95% confidential interval (CI): (2.571, 44.662)] and drinking alcohol [OR = 4.474, 95% CI: (1.019, 19.642)] were risk factors for biliary obstruction patients with C. sinensis infections, while living in city [OR = 0.128, 95% CI: (0.038, 0.435)] was a protective factor. Additionally, in biliary obstruction patients with C. sinensis infections, lower total bilirubin (Z = -2.566, P <0.05) and direct bilirubin (Z = -3.454, P <0.05), higher indirect bilirubin (Z = -3.821, P < 0.05), thickening of the bile duct wall and dilatation of the intrahepatic bile duct were detected. CONCLUSIONS: A comprehensive diagnosis requires to be made based on clinical symptoms, laboratory and imaging examinations, in order to improve the diagnosis of biliary obstruction patients with C. sinensis infections.

Current status of Clonorchis sinensis and clonorchiasis in Korea: epidemiological perspectives integrating the data from human and intermediate hosts.

Clonorchis sinensis is a carcinogenic liver fluke that causes clonorchiasis in humans. Clonorchiasis is prevalent in East Asian countries, and approximately 15­20 million individuals are estimated to be infected with this fluke globally. This review highlights the current status of C. sinensis and clonorchiasis in Korea from the epidemiological perspective involving the analysis of humans and intermediate hosts. Despite the recent decline in C. sinensis infection rate in Korea, C. sinensis infections remain endemic in 5 major river basins (Han-gang, Geum-gang, Seomjin-gang, Yeongsan-gang and Nakdong-gang; gang means river) with a high incidence of cholangiocarcinoma. A noticeable pattern involves increasing mild infections among patients diagnosed positive for C. sinensis eggs. The infection rate of C. sinensis metacercariae in the second intermediate host, freshwater fish, is also maintained at a substantial level. Thus, the One Health approach integrating different sectors and disciplines is recommended to accelerate and sustain control of C. sinensis, thereby leading to successful eradication. Health promotion via information dissemination and health education should be extended to prevent the consumption of raw freshwater fish by residents living in high-risk areas.

Multiple biochemical indices and metabolomics of Clonorchis sinensis provide a novel interpretation of biomarkers.

BACKGROUND: Clonorchiasis, an infectious disease caused by the liver fluke Clonorchis sinensis, may lead to the development of liver and gallbladder diseases, and even cholangiocarcinoma (CCA). However, the pathogenesis, host-pathogen interaction, and diagnostic markers for clonorchiasis remain unclear. METHODS: Eighteen rabbits were randomly divided into control group (n = 9) and C. sinensis-infected group (n = 9), and their plasma samples were collected at 7, 14, 28, and 63 days post-infection (dpi). Biochemical indices and metabolites in different infection periods were detected. A non-targeted ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) approach was employed to investigate the metabolic profiles of plasma in rabbits, and related metabolic pathways of differential metabolites and correlation between candidate biochemical indices and differential metabolites were analyzed. Finally, the candidate biomarkers were verified with human samples using a targeted metabolomics method. RESULTS: The result of biochemical indices indicated C. sinensis infection would affect the liver function biochemical indices, especially alanine aminotransferase, aspartate transaminase (AST), glutamyl transpeptidase (GGT), total bile acid, high-density lipoprotein, and cholinesterase. The metabonomic results showed that 58, 212, 23, and 21 differential metabolites were identified in different phases of the infection. Multivariate statistical analysis of differential metabolites revealed distinct metabolic signatures during different phases of infection, with most of these signatures being observed at 14 dpi, which mainly influences the amino acid metabolisms. For metabolites and biochemical indices, AST, GGT, hypoxanthine, L-pipecolic acid, and D-glucuronate represented potential noninvasive biomarkers for the diagnosis of C. sinensis (P < 0.05 and AUC > 0.8). Furthermore, GGT and D-glucuronate levels were positively correlated with the infection (r(28) = 0.98, P < 0.0001) and showed excellent diagnostic performance (AUC = 0.972; 95% confidence interval, 0.921 to 1.000). CONCLUSIONS: The present results provide new insights into plasma metabolic changes in rabbits during C. sinensis infection, and the potential biomarker may be used for developing an effective method to diagnose clonorchiasis in the future.

Identification of Myoferlin, a Potential Serodiagnostic Antigen of Clonorchiasis, via Immunoproteomic Analysis of Sera From Different Infection Periods and Excretory-Secretory Products of Clonorchis sinensis.

Clonorchiasis, which is caused by Clonorchis sinensis, is an important foodborne disease worldwide. The excretory-secretory products (ESPs) of C. sinensis play important roles in host-parasite interactions by acting as causative agents. In the present study, the ESPs and sera positive for C. sinensis were collected to identify proteins specific to the sera of C. sinensis (i.e., proteins that do not cross-react with Fasciola hepatica and Schistosoma japonicum) at different infection periods. Briefly, white Japanese rabbits were artificially infected with C. sinensis, and their sera were collected at 7 days post-infection (dpi), 14 dpi, 35 dpi, and 77 dpi. To identify the specific proteins in C. sinensis, a co-immunoprecipitation (Co-IP) assay was conducted using shotgun liquid chromatography tandem-mass spectrometry (LC-MS/MS) to pull down the sera roots of C. sinensis, F. hepatica, and S. japonicum. For the annotated proteins, 32, 18, 39, and 35 proteins specific to C. sinensis were pulled down by the infected sera at 7, 14, 35, and 77 dpi, respectively. Three proteins, Dynein light chain-1, Dynein light chain-2 and Myoferlin were detected in all infection periods. Of these proteins, myoferlin is known to be overexpressed in several human cancers and could be a promising biomarker and therapeutic target for cancer cases. Accordingly, this protein was selected for further studies. To achieve a better expression, myoferlin was truncated into two parts, Myof1 and Myof2 (1,500 bp and 810 bp), based on the antigenic epitopes provided by bioinformatics. The estimated molecular weight of the recombinant proteins was 57.3 ku (Myof1) and 31.3 ku (Myof2). Further, both Myof1 and Myof2 could be probed by the sera from rabbits infected with C. sinensis. No cross-reaction occurred with the positive sera of S. japonica, F. hepatica, and negative controls. Such findings indicate that myoferlin may be an important diagnostic antigen present in the ESPs. Overall, the present study provides new insights into proteomic changes between ESPs and hosts in different infection periods by LC-MS/MS. Moreover, myoferlin, as a biomarker, may be used to develop an objective method for future diagnosis of clonorchiasis.

A CLINICAL CASE OF COEXISTENT PARASITIC LIVER AND GASTRIC LESIONS CAUSED BY CLONORCHIASIS.

Clonorchiasis is a parasitic disease caused by Clonorchis sinensis. Parasite colonies can develop not only in the bile and pancreatic ducts but also in the gastric wall. This is confirmed by the described clinical case of perforated gastric ulcer, the morphological study of which revealed parasite colonies in the wall of the organ.

Case Report: Clonorchis sinensis Infection Associated with Eosinophilic Pneumonia.

Clonorchis sinensis, a trematode prevalent in East Asia, causes hepatobiliary infection. Exposure typically occurs through ingestion of raw or undercooked fish containing the encysted larval form of the parasite. Extrahepatobiliary disease has not commonly been described. In this case report, we describe an unusual case of C. sinensis infection associated with eosinophilic pneumonia. A middle-aged man from China presented with subacute cough and was found to have a bilateral diffuse eosinophilic pneumonia with associated peripheral eosinophilia. Stool microscopy revealed C. sinensis eggs, and the patient improved after treatment with prednisone and praziquantel. Pulmonary clonorchiasis should be considered in patients with eosinophilic pneumonia from areas highly endemic for this pathogen.

Severe hepatobiliary morbidity is associated with Clonorchis sinensis infection: The evidence from a cross-sectional community study.

Clonorchis sinensis infection is highly prevalent in Asia. Diverse hepatobiliary morbidity has been documented for C. sinensis infection. This study aimed to assess the association between C. sinensis infection and hepatobiliary morbidity, taking into consideration of the control, confounders and infection intensity. A cross-sectional community survey was implemented in Hengxian county, southeastern China. Helminth infections were detected by fecal examination. Physical examination and abdominal ultrasonography were then conducted. After excluding confounding effects from gender, age and alcohol drinking, quantitative association between C. sinensis infection and hepatobiliary morbidity was assessed, and the effect from infection intensity was also evaluated, through adjusted odds ratio (aOR) and 95% confidence intervals (95% CI). 696 villagers older than 10 years were enrolled. The prevalence and infection intensity of C. sinensis were higher in male, elder people and the individuals consuming alcohol. Light C. sinensis infection was associated with the increase of diarrhoea (aOR: 2.2, 95% CI: 1.1-4.5). C. sinensis infection was associated with the increase of fatty liver (aOR: 2.7, 95% CI: 1.4-5.2), and the effect was similar in different infection intensities. Moderate C. sinensis infection was associated with the increase of gallbladder stone (aOR: 3.0, 95% CI: 1.1-8.6), while moderate and heavy infections with the increase of intrahepatic bile duct dilatation (aOR: 2.2, 95% CI: 1.0-4.9 and aOR: 4.3, 95% CI: 1.9-9.9, respectively). C. sinensis infection had an effect on the development of periductal fibrosis (aOR: 3.2, 95% CI: 2.1-4.9), which showed increasing trend by infection intensity. The length and width of gallbladder in those with C. sinensis infection were enlarged, especially in those over 30 years old. C. sinensis infection is significantly associated with hepatobiliary morbidity. The occurrence of some morbidity was strongly related to the infection intensity. Awareness on harm of clonorchiasis should be raised both for policy-makers and villagers to adopt effective interventions.

[Prevalence of Clonorchis sinensis infections in freshwater fish in mainland China: A meta-analysis].

OBJECTIVE: To understand the real prevalence of Clonorchis sinensis infections in the freshwater fish in mainland China, so as to provide insights into clonorchiasis control and detection of freshwater fish. METHODS: All literatures reporting the prevalence of C. sinensis infections in the freshwater fish, the second intermediate host of the parasite, were jointly retrieved in Chinese and English electronic databases from January 1, 2010 to December 31, 2020, including Wanfang Data, CNKI, PubMed, Web of Science, Embase and Cochrane Library. All studies were screened based on inclusion and exclusion criteria, and the quality of all enrolled literatures was evaluated. The pooled prevalence of C. sinensis infections in freshwater fish and its 95% confidence interval (CI) were estimated using the software Stata version 15.0, and subgroup analyses were performed to investigate the region-, season- and sample source-specific pooled prevalence of C. sinensis infections in freshwater fish. In addition, the sensitivity and publication bias of all included studies were analyzed. RESULTS: A total of 40 eligible literatures were included in this study, including 37 Chinese literatures and 3 English literatures, and there were 10 high-quality literatures, 27 moderate-quality literatures and 3 low-quality literatures. A total of 53 species containing 37 959 freshwater fish were reported in these 40 studies, and 73.58% (39/53) of freshwater fish species were identified with C. sinensis infections. Meta-analysis showed 23.5% [95% CI: (0.19, 0.28)] pooled prevalence of C. sinensis infections in freshwater fish in mainland China, and subgroup analyses higher prevalence of C. sinensis infections in freshwater fish in northeastern China [35.7%, 95% CI: (0.22, 0.50)] than in central [25.9%, 95% CI: (0.04, 0.48)] and southern China [20.6%, 95% CI: (0.09, 0.32)], higher prevalence of C. sinensis infections in freshwater fish sampled in spring [44.1%, 95% CI: (0.35, 0.53)] than in autumn [6.7%, 95% CI: (0.05, 0.08)] and summer [3.3%, 95% CI: (-0.01, 0.07)], and higher prevalence of C. sinensis infections in freshwater fish sampled from natural water [25.2%, 95% CI: (0.17, 0.33)] than from retail trades [22.2%, 95% CI: (0.17, 0.28)] and breeding chain [12.3%, 95% CI: (0.03, 0.22)]. However, all included studies had a publication bias with a low sensitivity. CONCLUSIONS: The prevalence of C. sinensis infections is high in freshwater fish in mainland China, and there are still challenges for clonorchiasis control. Reinforcement of health education, diagnostics development and food safety supervision is recommended in future clonorchiasis control programs.

Serodiagnostic antigens of Clonorchis sinensis identified and evaluated by high-throughput proteogenomics.

Clonorchiasis caused by Clonorchis sinensis is endemic in East Asia; approximately 15 million people have been infected thus far. To diagnose the infection, serodiagnostic tests with excellent functionality should be performed. First, 607 expressed sequence tags encoding polypeptides with a secretory signal were expressed into recombinant proteins using an in vitro translation system. By protein array-based screening using C. sinensis-infected sera, 18 antigen candidate proteins were selected and assayed for cross-reactivity against Opisthorchis viverrini-infected sera. Of the six antigenic proteins selected, four were synthesized on large scale in vitro and evaluated for antigenicity against the flukes-infected human sera using ELISA. CsAg17 antigen showed the highest sensitivity (77.1%) and specificity (71.2%). The sensitivity and specificity of the bacterially produced CsAg17-28GST fusion antigen was similar to those of CsAg17 antigen. CsAg17 antigen can be used to develop point-of-care serodiagnostic tests for clonorchiasis.

Bile Ductal Transcriptome Identifies Key Pathways and Hub Genes in Clonorchis sinensis-Infected Sprague-Dawley Rats.

Clonorchis sinensis is a food-borne trematode that infects more than 15 million people. The liver fluke causes clonorchiasis and chronical cholangitis, and promotes cholangiocarcinoma. The underlying molecular pathogenesis occurring in the bile duct by the infection is little known. In this study, transcriptome profile in the bile ducts infected with C. sinensis were analyzed using microarray methods. Differentially expressed genes (DEGs) were 1,563 and 1,457 at 2 and 4 weeks after infection. Majority of the DEGs were temporally dysregulated at 2 weeks, but 519 DEGs showed monotonically changing expression patterns that formed seven distinct expression profiles. Protein-protein interaction (PPI) analysis of the DEG products revealed 5 sub-networks and 10 key hub proteins while weighted co-expression network analysis (WGCNA)-derived gene-gene interaction exhibited 16 co-expression modules and 13 key hub genes. The DEGs were significantly enriched in 16 Kyoto Encyclopedia of Genes and Genomes pathways, which were related to original systems, cellular process, environmental information processing, and human diseases. This study uncovered a global picture of gene expression profiles in the bile ducts infected with C. sinensis, and provided a set of potent predictive biomarkers for early diagnosis of clonorchiasis.

Association between renal function and co-infection with Clonorchis sinensis and Helicobacter pylori: a cross-sectional study.

BACKGROUND: Studies have shown that liver fluke infections may be associated with kidney injury and that Helicobacter pylori (Hp) may be involved in the pathogenesis of kidney diseases. However, no studies have reported the relationship between co-infection with Clonorchis sinensis (Cs) and Hp and renal function. The aim of this study was to examine the relationship between co-infection with Cs and Hp and estimated glomerular filtration rate (eGFR) in a general population, and gender-related differences were also investigated. METHODS: In the cross-sectional study, 4122 subjects from the Health Examination Center of Guangdong Provincial Hospital of Integrated Traditional Chinese and Western Medicine from January 2017 to December 2018 were enrolled. All participants underwent stool examination for the diagnosis of Cs infection and 13C-urea breath test (UBT) for the diagnosis of Hp infection. Participants were categorized into four groups: (1) co-infection with Cs and Hp group comprising 207 cases (Hp(+) + Cs(+) group), (2) Cs infection group comprising 1392 cases (Hp(-) + Cs(+)group), (3) Hp infection group comprising 275 cases (Hp(+) + Cs(-) group), and (4) non-infection group comprising 2248 cases (Hp(-) + Cs(-) group). Multiple linear regression analysis was performed to evaluate the relationship between co-infection with Cs and Hp and eGFR. RESULTS: Hp infection without Cs infection was present in 6.67% (275/4122) of subjects, while Cs infection without Hp infection was present in 33.77% (1392/4122) of subjects. Co-infection with Hp and Cs were present in 5.02% (207/4122) of subjects. Median age of the participants was 43 years (IQR 35-51). Most of the participants were male (2955/4122, 71.69%). Median eGFR was 96.61 ml/min/1.73 m2 (IQR 85.05-106.24). Co-infection with Cs and Hp was negatively associated with eGFR after full adjusting (ß = - 1.89, 95% CI: - 3.33 to - 0.45, p = 0.01). The relationship remained significant in females (ß = - 9.37, 95% CI: - 11.60 to - 7.1, p < 0.001), but not in males. CONCLUSION: Our findings suggest that co-infection with Cs and Hp may be associated with reduced renal function in females, but not in males.