Dataset Information

Detection of the circulating antigen 14-3-3 protein of Schistosoma japonicum by time-resolved fluoroimmunoassay in rabbits.

ABSTRACT: BACKGROUND: Schistosomiasis remains a major public health concern that afflicts millions of people worldwide. Low levels of Schistosoma infection require more sensitive diagnostic methods. In this study, a time-resolved fluoroimmunoassay (TRFIA) was developed for detecting the signal transduction protein 14-3-3, a circulating antigen of Schistosoma japonicum. RESULTS: The detection limit of 14-3-3-TRFIA was 0.78 ng/ml, with a linear measurement range from 0.78 to 800 ng/ml. The average intra-assay and inter-assay variability of this TRFIA was 8.9% and 12.2% respectively, and the mean recovery rate ranged from 92.1% to 115.5%. Within the first 21 days post-infection in rabbits, the positive rates of the 14-3-3-TRFIA were distinctly higher compared to ELISA. All these findings illustrate that 14-3-3-TRFIA has a higher detection efficacy and is a good early diagnostic method for active Schistosoma infection. CONCLUSIONS: A sandwich TRFIA for detecting the circulating antigen 14-3-3 of S. japonicum has been developed, and has demonstrated to be a good potential diagnostic method for schistosomiasis.

SUBMITTER: Qian CY

PROVIDER: S-EPMC3115898 | biostudies-other | 2011

REPOSITORIES: biostudies-other

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Publications

Detection of the circulating antigen 14-3-3 protein of Schistosoma japonicum by time-resolved fluoroimmunoassay in rabbits.

Qian Chun-Yan CY Huang Biao B Yu Chuan-Xin CX Zhang Jue J Yin Xu-Ren XR Wang Jie J Song Li-Jun LJ Zhang Wei W Ke Xue-Dan XD

Parasites & vectors 20110528

<h4>Background</h4>Schistosomiasis remains a major public health concern that afflicts millions of people worldwide. Low levels of Schistosoma infection require more sensitive diagnostic methods. In this study, a time-resolved fluoroimmunoassay (TRFIA) was developed for detecting the signal transduction protein 14-3-3, a circulating antigen of Schistosoma japonicum.<h4>Results</h4>The detection limit of 14-3-3-TRFIA was 0.78 ng/ml, with a linear measurement range from 0.78 to 800 ng/ml. The aver ...[more]

PMID: 21619661

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Project description:Dioecious flatworms of the genus Schistosoma causes schistosomiasis, which is a major public health problem in developing countries. Acquiring detailed knowledge of schistosome-host interactions may aid in the development of novel strategies for schistosomiasis control. MicroRNAs (miRNAs) are involved in processes such as development, cell proliferation, metabolism, and signal transduction. Circulating miRNAs not only serve as a novel class of biomarkers of many diseases but also regulate target gene expression in recipient cells, which are similar to hormones.In the present study, we used miRNA microarrays to determine the profile of circulating miRNAs associated with S. japonicum infection of mice. The biological functions of the altered levels of miRNAs and their target genes were predicted using bioinformatics. Expression levels of selected miRNAs and their target genes were further analyzed by quantitative RT-PCR.Our study identified 294 and 189 miRNAs in infected mice that were expressed in two independent experiments at levels???2-fold higher or???0.5-fold lower, respectively, compared with uninfected mice. Thirty-six of the same miRNAs were detected in these analyses. Moreover, pathway analyses indicated that most of these miRNAs are putatively involved in signaling pathways associated with pathogenesis, such as Wnt and MAPK signaling. Further, we show an inverse correlation between the circulating levels of these miRNAs and their target genes, suggesting that changes in miRNA expression may cause aberrant expression of genes such as Creb1 and Caspase-3 in mice infected with S. japonicum.Our study shows significant differences in the levels of circulating miRNAs between S. japonicum infected mice and uninfected mice. In particular, the altered levels of miR-706 and miR-134-5p were associated with altered levels of expression of the Caspase-3 and Creb1 genes, respectively, suggesting that circulating miRNAs may serve as important mediators of the pathology of hepatic schistosomiasis. Additionally, our results are expected to provide new insights for further understanding the mechanisms of schistosome-host interaction that may facilitate in the development of novel interventions for alleviating the symptom of S. japonicum infection as well as for preventing and treating schistosomiasis.

Project description:Schistosomiasis japonica is a common zoonosis. Domestic animals are the primary source of infection and play an important role in disease transmission. The prevalence and infectivity of this disease in domestic animals in China have significantly decreased and, for this reason, diagnostics with a higher sensitivity have become increasingly necessary. It was reported that polymerase chain reaction (PCR)-based methods could be used to detect schistosome infection in humans and animals and presented a high sensitivity and specificity. The present study aimed to develop a PCR-based method for detection of Schistosoma japonicum infection in domestic animals.A specific nested-PCR assay was developed to detect S. japonicum infection in domestic animals via amplification of a 231-bp DNA fragment of retrotransposon SjR2. The developed assay was first used in sera and dry blood filter paper (DBFP) from goats and buffaloes at different time points of infection. Then, 78 DBFPs from 39 artificially-infected bovines at 14 and 28 days post-infection and 42 DBFPs from schistosome-negative bovines from the city of Huangshan in the Anhui province were used to evaluate the diagnostic validity. Furthermore, this assay was used to detect S. japonicum infection in domestic animals in Dongzhi and Wangjiang counties.The expected PCR product was detected in eggs and adult worms of S. japonicum and blood samples from S. japonicum-infected goats and water buffaloes, but not from Fasciola and Haemonchus contortus worms. The nested-PCR assay could detect the target S. japonicum DNA in DBFPs from goats and buffaloes after day 3 post-infection. The sensitivity in buffaloes at 14 and 28 days post-infection was 92.30% (36/39) and 100% (39/39), respectively. The specificity was 97.60% (41/42). The positivity rates in Dongzhi and Wangjiang counties were 6.00% and 8.00% in bovines and 22.00% and 16.67% in goats, respectively. The positivity rates in goats in both counties were higher than those in bovines with a significant difference in Dongzhi County but not in Wangjiang County (P?<?0.05 and P?=?0.23, respectively).Our results suggest that the developed nested-PCR assay may be used for the diagnosis of S. japonicum infection in domestic animals, and the control of S. japonicum infection in goats should be paid more attention.

Project description:Background Elimination and control of Schistosoma japonicum, the most virulent of the schistosomiasis-causing blood flukes, requires the development of sensitive and specific diagnostic tools capable of providing an accurate measurement of the infection prevalence in endemic areas. Typically, detection of S. japonicum has occurred using the Kato-Katz technique, but this methodology, which requires skilled microscopists, has been shown to radically underestimate levels of infection. With the ever-improving capabilities of next-generation sequencing and bioinformatic analysis tools, identification of satellite sequences and other highly repetitive genomic elements for use as real-time PCR diagnostic targets is becoming increasingly common. Assays developed using these targets have the ability to improve the sensitivity and specificity of results for epidemiological studies that can in turn be used to inform mass drug administration and programmatic decision making. Methodology/Principal findings Utilizing Tandem Repeat Analyzer (TAREAN) and RepeatExplorer2, a cluster-based analysis of the S. japonicum genome was performed and a tandemly arranged genomic repeat, which we named SjTR1 (SchistosomajaponicumTandem Repeat 1), was selected as the target for a real-time PCR diagnostic assay. Based on these analyses, a primer/probe set was designed and the assay was optimized. The resulting real-time PCR test was shown to reliably detect as little as 200 ag of S. japonicum genomic DNA and as little as 1 egg per gram of human stool. Based on these results, the index assay reported in this manuscript is more sensitive than previously published real-time PCR assays for the detection of S. japonicum. Conclusions/Significance The extremely sensitive and specific diagnostic assay described in this manuscript will facilitate the accurate detection of S. japonicum, particularly in regions with low levels of endemicity. This assay will be useful in providing data to inform programmatic decision makers, aiding disease control and elimination efforts. Author summary Schistosomiasis is a Neglected Tropical Disease (NTD) estimated to infect more than 230 million people worldwide. Of the various species of schistosomes that cause disease in humans, Schistosoma japonicum is considered the most virulent. As such, this pathogen presents a crucial public health threat. Typically, diagnosis of S. japonicum has been performed via the Kato-Katz technique which has been shown to dramatically underestimate the burden of infection, resulting in a need for improved detection strategies. To address this need, we have employed bioinformatic tools in order to identify a tandemly arranged, highly repetitive, DNA sequence, SjTR1, in the S. japonicum genome. Utilizing this sequence as a real-time PCR assay target, we have developed a sensitive and specific assay for the detection of S. japonicum DNA. Employment of this assay in field settings will facilitate the accurate detection of S. japonicum and provide guidance capable of informing mass drug administration efforts targeting elimination.

Dataset Information

Detection of the circulating antigen 14-3-3 protein of Schistosoma japonicum by time-resolved fluoroimmunoassay in rabbits.

Publications

Detection of the circulating antigen 14-3-3 protein of Schistosoma japonicum by time-resolved fluoroimmunoassay in rabbits.

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