{"title":"巢式-qPCR-高分辨熔融(HRM)技术在伊朗寄生虫中的应用","authors":"","doi":"10.1007/s11686-024-00821-2","DOIUrl":null,"url":null,"abstract":"<h3>Abstract</h3> <span> <h3>Purpose</h3> <p><em>Strongyloides stercoralis</em> is a parasite with special characteristics presenting it as a unique nematode. Iran is an endemic area for <em>S.</em> <em>stercoralis.</em> In this study, nested-qPCR-high resolution melting (HRM) technology was applied on some human isolates of <em>S.</em> <em>stercoralis</em> from this country by focusing on evolutionary genetics analysis.</p> </span> <span> <h3>Methods</h3> <p>Twelve human isolates of <em>S. stercoralis</em> were collected from four endemic provinces of Iran. Genomic DNA was extracted from a single filariform larva for every isolate. Using specific primers targeting partial regions in <em>cox</em>1 gene, nested-qPCR-HRM was performed and melting-curve profiles were analyzed alongside the evaluation of genetic proximity and phylogenetic analysis using MEGA7 and DnaSP5 software.</p> </span> <span> <h3>Results</h3> <p>The melting temperature (T<sub>m</sub>) values of the isolates were 77.9 °C−78.3 °C. All isolates from Guilan, Mazandaran, and Khouzestan Provinces shared T<sub>m</sub> values of 78.2 °C to 78.3 °C, while the isolates from Hormozgan Province showed T<sub>m</sub> values of 77.9 °C, 78.0 °C, and 78.1 °C. The phylogenetic tree illustrated that the sequences of the current study included nine haplotypes. Tajima’s D index analyses showed that <em>cox</em>1 gene in <em>S. stercoralis</em> isolates was negative (Tajima’s <em>D</em> = <sub>−</sub> 0.27).</p> </span> <span> <h3>Conclusion</h3> <p>The isolates were divided into five temperature groups. Although HRM assay compared to PCR sequencing identified more limited genetic changes, it revealed that the mean of T<sub>m</sub> of the isolates from Hormozgan Province was lower than those of other provinces and represented specific haplotypes for this geographical region on the phylogenetic tree.</p> </span>","PeriodicalId":6932,"journal":{"name":"Acta Parasitologica","volume":null,"pages":null},"PeriodicalIF":1.2000,"publicationDate":"2024-03-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"Application of Nested-qPCR-High Resolution Melting (HRM) Technology on Strongyloides stercoralis Isolates from Iran\",\"authors\":\"\",\"doi\":\"10.1007/s11686-024-00821-2\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<h3>Abstract</h3> <span> <h3>Purpose</h3> <p><em>Strongyloides stercoralis</em> is a parasite with special characteristics presenting it as a unique nematode. Iran is an endemic area for <em>S.</em> <em>stercoralis.</em> In this study, nested-qPCR-high resolution melting (HRM) technology was applied on some human isolates of <em>S.</em> <em>stercoralis</em> from this country by focusing on evolutionary genetics analysis.</p> </span> <span> <h3>Methods</h3> <p>Twelve human isolates of <em>S. stercoralis</em> were collected from four endemic provinces of Iran. Genomic DNA was extracted from a single filariform larva for every isolate. Using specific primers targeting partial regions in <em>cox</em>1 gene, nested-qPCR-HRM was performed and melting-curve profiles were analyzed alongside the evaluation of genetic proximity and phylogenetic analysis using MEGA7 and DnaSP5 software.</p> </span> <span> <h3>Results</h3> <p>The melting temperature (T<sub>m</sub>) values of the isolates were 77.9 °C−78.3 °C. All isolates from Guilan, Mazandaran, and Khouzestan Provinces shared T<sub>m</sub> values of 78.2 °C to 78.3 °C, while the isolates from Hormozgan Province showed T<sub>m</sub> values of 77.9 °C, 78.0 °C, and 78.1 °C. The phylogenetic tree illustrated that the sequences of the current study included nine haplotypes. Tajima’s D index analyses showed that <em>cox</em>1 gene in <em>S. stercoralis</em> isolates was negative (Tajima’s <em>D</em> = <sub>−</sub> 0.27).</p> </span> <span> <h3>Conclusion</h3> <p>The isolates were divided into five temperature groups. Although HRM assay compared to PCR sequencing identified more limited genetic changes, it revealed that the mean of T<sub>m</sub> of the isolates from Hormozgan Province was lower than those of other provinces and represented specific haplotypes for this geographical region on the phylogenetic tree.</p> </span>\",\"PeriodicalId\":6932,\"journal\":{\"name\":\"Acta Parasitologica\",\"volume\":null,\"pages\":null},\"PeriodicalIF\":1.2000,\"publicationDate\":\"2024-03-12\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Acta Parasitologica\",\"FirstCategoryId\":\"3\",\"ListUrlMain\":\"https://doi.org/10.1007/s11686-024-00821-2\",\"RegionNum\":3,\"RegionCategory\":\"农林科学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q4\",\"JCRName\":\"PARASITOLOGY\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Acta Parasitologica","FirstCategoryId":"3","ListUrlMain":"https://doi.org/10.1007/s11686-024-00821-2","RegionNum":3,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q4","JCRName":"PARASITOLOGY","Score":null,"Total":0}
Application of Nested-qPCR-High Resolution Melting (HRM) Technology on Strongyloides stercoralis Isolates from Iran
Abstract
Purpose
Strongyloides stercoralis is a parasite with special characteristics presenting it as a unique nematode. Iran is an endemic area for S.stercoralis. In this study, nested-qPCR-high resolution melting (HRM) technology was applied on some human isolates of S.stercoralis from this country by focusing on evolutionary genetics analysis.
Methods
Twelve human isolates of S. stercoralis were collected from four endemic provinces of Iran. Genomic DNA was extracted from a single filariform larva for every isolate. Using specific primers targeting partial regions in cox1 gene, nested-qPCR-HRM was performed and melting-curve profiles were analyzed alongside the evaluation of genetic proximity and phylogenetic analysis using MEGA7 and DnaSP5 software.
Results
The melting temperature (Tm) values of the isolates were 77.9 °C−78.3 °C. All isolates from Guilan, Mazandaran, and Khouzestan Provinces shared Tm values of 78.2 °C to 78.3 °C, while the isolates from Hormozgan Province showed Tm values of 77.9 °C, 78.0 °C, and 78.1 °C. The phylogenetic tree illustrated that the sequences of the current study included nine haplotypes. Tajima’s D index analyses showed that cox1 gene in S. stercoralis isolates was negative (Tajima’s D = − 0.27).
Conclusion
The isolates were divided into five temperature groups. Although HRM assay compared to PCR sequencing identified more limited genetic changes, it revealed that the mean of Tm of the isolates from Hormozgan Province was lower than those of other provinces and represented specific haplotypes for this geographical region on the phylogenetic tree.
期刊介绍:
Acta Parasitologica is an international journal covering the latest advances in the subject.
Acta Parasitologica publishes original papers on all aspects of parasitology and host-parasite relationships, including the latest discoveries in biochemical and molecular biology of parasites, their physiology, morphology, taxonomy and ecology, as well as original research papers on immunology, pathology, and epidemiology of parasitic diseases in the context of medical, veterinary and biological sciences. The journal also publishes short research notes, invited review articles, book reviews.
The journal was founded in 1953 as "Acta Parasitologica Polonica" by the Polish Parasitological Society and since 1954 has been published by W. Stefanski Institute of Parasitology of the Polish Academy of Sciences in Warsaw. Since 1992 in has appeared as Acta Parasitologica in four issues per year.