Construction and application of the RPA-LFD rapid detection method for <italic style="font-style: italic">Glugea plecoglossi</italic>

Jiaxue SONG; Ruixin FENG; Yunfei PANG; Qingyue XU; Dong ZHENG; Yunji XIU; Shun ZHOU

doi:10.1007/s00343-024-4173-y

Your Location：

Home >

Browse articles >

Construction and application of the RPA-LFD rapid detection method for Glugea plecoglossi

Aquaculture and Fisheries | Updated：2025-10-16

- Construction and application of the RPA-LFD rapid detection method for Glugea plecoglossi
- Construction and application of the RPA-LFD rapid detection method for Glugea plecoglossi
- 海洋湖沼学报(英文) 2025年43卷第5期页码：1647-1653
- Affiliations：
  
  School of Marine Science and Engineering, Qingdao Agricultural University, Qingdao 266109, China
- Author bio：
  
  yunji16@163.com
  zhoushun@qau.edu.cn
- Funds：
  
  the Visiting and Training Foundation of Teachers in Ordinary Undergraduate Universities of Shandong Province, the Qingdao Agricultural University Doctoral Start-Up Fund(6631122030);the Advanced Talents Foundation of QAU(6651118016);the Fish Innovation Team of Shandong Agriculture Research System(SDAIT-12-06);the Shandong Engineering Research Center for Prevention and Control of Aquatic Animal Disease, the “First Class Fishery Discipline” Program((2020)3);of Shandong Province, and the Key RD Program (Soft Science Project) of Shandong Province, China(2023RKY06004)
- DOI：10.1007/s00343-024-4173-y
  中图分类号：
- 收稿：2024-06-27，
  
  录用：2024-09-02，
  
  网络首发：2024-10-16，
  
  纸质出版：2025-09-01
- Accepted：
Scan QR Code
Construction and application of the RPA-LFD rapid detection method for Glugea plecoglossi[J]. 海洋湖沼学报(英文), 2025,43(5):1647-1653.

SONG Jiaxue,FENG Ruixin,PANG Yunfei,et al.Construction and application of the RPA-LFD rapid detection method for ,Glugea plecoglossi[J].Journal of Oceanology and Limnology,2025,43(05):1647-1653.
Construction and application of the RPA-LFD rapid detection method for Glugea plecoglossi[J]. 海洋湖沼学报(英文), 2025,43(5):1647-1653. DOI： 10.1007/s00343-024-4173-y.

SONG Jiaxue,FENG Ruixin,PANG Yunfei,et al.Construction and application of the RPA-LFD rapid detection method for ,Glugea plecoglossi[J].Journal of Oceanology and Limnology,2025,43(05):1647-1653. DOI： 10.1007/s00343-024-4173-y.

摘要

Abstract

Glugea plecoglossi

a microsporidia of the

Glugea

genus

can cause an infamous disease

Plecoglossus

altivelis

in East Asia

resulting in heavy economic losses. At present

the main diagnostic methods for this disease include microscopy examination

quantitative real-time PCR

and loop-mediated isothermal amplification-lateral flow dipstick (LAMP-LFD). In this study

a recombinase polymerase amplification-lateral flow dipstick (RPA-LFD) method

targeting the

beta

tubulin

gene

was developed to detect

plecoglossi

three sets of primers and probes were designed and screened

after which the initial reaction system was established. The RPA-LFD method for

plecoglossi

could complete nucleic acid amplification at 39 °C for 10 min

after which the amplification product was dropped on the LFD strip

and the results could then be observed within 5 min. A specificity assay revealed that there was no cross-reactivity with other protozoa except

plecoglossi

. A sensitivity assay revealed that the detection limit was 9.38×10

ng/μL

which was more sensitive than that of conventional PCR. Compared with conventional detection methods

the novel RPA-LFD method has the advantages of simple operation

short operation time

high sensitivity

and high specificity for

plecoglossi

detection

indicating its potential use in rapid field detection of

plecoglossi

关键词

Keywords

references

An X , Zhao Y , Cui J et al . 2021 . Recombinase polymerase amplification lateral flow dipstick (RPA-LF) detection of Babesia orientalis in water buffalo ( Bubalus babalis , Linnaeus , 1758) . Veterinary Parasitology , 296 , https://doi.org/10.1016/j.vetpar.2021.109479 https://doi.org/10.1016/j.vetpar.2021.109479 .

Ashley P J . 2007 . Fish welfare: current issues in aquaculture . Applied Animal Behaviour Science , 104 ( 3-4 ): 199 - 235 , https://doi.org/10.1016/j.applanim.2006.09.001 https://doi.org/10.1016/j.applanim.2006.09.001 .

Boyle D S , McNerney R , Low H T et al . 2014 . Rapid detection of Mycobacterium tuberculosis by recombinase polymerase amplification . PLoS One , 9 ( 8 ): e 103091 , https://doi.org/10.1371/journal.pone.0103091 https://doi.org/10.1371/journal.pone.0103091 .

Casal G , Rocha S , Costa G et al . 2016 . Ultrastructural and molecular characterization of Glugea serranus n. sp., a microsporidian infecting the blacktail comber, Serranus atricauda (Teleostei: Serranidae), in the Madeira Archipelago (Portugal) . Parasitology Research , 115 ( 10 ): 3963 - 3972 , https://doi.org/10.1007/s00436-016-5162-7 https://doi.org/10.1007/s00436-016-5162-7 .

Cong F , Zeng F W , Wu M L et al . 2020 . Development of a real-time reverse transcription recombinase polymerase amplification assay for rapid detection of spring viremia of carp virus. Molecular and Cellular Probes , 50 : 101494 , https://doi.org/10.1016/j.mcp.2019.101494 https://doi.org/10.1016/j.mcp.2019.101494 .

Crannell Z A , Cabada M M , Castellanos-Gonzalez A et al . 2015 . Recombinase polymerase amplification-based assay to diagnose Giardia in stool samples . The American Journal of Tropical Medicine and Hygiene , 92 ( 3 ): 583 - 587 , https://doi.org/10.4269/ajtmh.14-0593 https://doi.org/10.4269/ajtmh.14-0593 .

Duc T H , Kinoshita I , Xuan N H et al . 2018 . Early life stages and habitats of ayu Plecoglossus altivelis (Temminck and Schlegel 1846) from two river-estuary systems in Vietnam . Asian Fisheries Science , 31 : 1 - 16 , https://doi.org/10.33997/j.afs.2018.31.1.001 https://doi.org/10.33997/j.afs.2018.31.1.001 .

Fan X X , Li L , Zhao Y G et al . 2020 . Clinical validation of two recombinase-based isothermal amplification assays (RPA/RAA) for the rapid detection of African swine fever virus. Frontiers in Microbiology , 11 : 1696 , https://doi.org/10.3389/fmicb.2020.01696 https://doi.org/10.3389/fmicb.2020.01696 .

Gao W F , Huang H L , Zhu P et al . 2018 . Recombinase polymerase amplification combined with lateral flow dipstick for equipment-free detecti on of Salmonella in shellfish . Bioprocess and Biosystems Engineering , 41 ( 5 ): 603 - 611 , https://doi.org/10.1007/s00449-018-1895-2 https://doi.org/10.1007/s00449-018-1895-2 .

Hata M , Otake T . 2019 . Downstream migration and mortality of larval Ayu ( Plecoglossus altivelis altivelis ) on the Sanriku coast, northern Japan . Environmental Biology of Fishes , 102 ( 10 ): 1311 - 1325 , https://doi.org/10.1007/s10641-019-00909-z https://doi.org/10.1007/s10641-019-00909-z .

He L , Zhou Y Q , Oosthuizen M C et al . 2009 . Loop-mediated isothermal amplification (LAMP) detection of Babesia orientalis in water buffalo ( Bubalus babalis , Linnaeus, 1758) in China . Veterinary Parasitology , 165 ( 1-2 ): 36 - 40 , https://doi.org/10.1016/j.vetpar.2009.06.036 https://doi.org/10.1016/j.vetpar.2009.06.036 .

Jaroenram W , Owens L . 2014 . Recombinase polymerase amplification combined with a lateral flow dipstick for discriminating between infectious Penaeus stylirostris densovirus and virus-related sequences in shrimp genome . Journal of Virological Methods , 208 : 144 - 151 , https://doi.org/10.1016/j.jviromet.2014.08.006 https://doi.org/10.1016/j.jviromet.2014.08.006 .

Jiang W , Ren Y , Han X et al . 2020 . Recombinase polymerase amplification-lateral flow (RPA-LF) assay combined with immunomagnetic separation for rapid visual detection of Vibrio parahaemolyticus in raw oysters . Analytical and Bioanalytical Chemistry , 412 ( 12 ): 2903 - 2914 , https://doi.org/10.1007/s00216-020-02532-9 https://doi.org/10.1007/s00216-020-02532-9 .

Joh S J , Kwon Y K , Kim M C et al . 2007 . Heterosporis anguillarum infections in farm cultured eels ( Anguilla japonica ) in Korea . Journal of Veterinary Science , 8 ( 2 ): 147 - 149 , https://doi.org/10.4142/jvs.2007.8.2.147 https://doi.org/10.4142/jvs.2007.8.2.147 .

Kang J W , Jin J L , Duan L J et al . 2020 . A loop-mediated isothermal amplification technique combined with a lateral flow dipstick for the detection of Glugea plecoglossi . Oceanologia et Limnologia Sinica , 51 ( 6 ): 1501 - 1512 , https://doi.org/10.11693/hyhz20200300076. https://doi.org/10.11693/hyhz20200300076. (in Chinese with English abstract)

Karthik L , Kumar G , Keswani T et al . 2014 . Protease inhibitors from marine actinobacteria as a potential source for antimalarial compound . PLoS One , 9 ( 3 ): e 90972 , https://doi.org/10.1371/journal.pone.0090972 https://doi.org/10.1371/journal.pone.0090972 .

Kralik P , Ricchi M . 2017 . A basic guide to real time PCR in microbial diagnostics: definitions, parameters, and everything. Frontiers in Microbiology , 8 : 108 , https://doi.org/10.3389/fmicb.2017.00108 https://doi.org/10.3389/fmicb.2017.00108 .

Lee S J , Yokoyama H , Ogawa K . 2004 . Modes of transmission of Glugea plecoglossi (Microspora) via the skin and digestive tract in an experimental infection model using rainbow trout, Oncorhynchus mykiss (Walbaum) . Journal of Fish Diseases , 27 ( 8 ): 435 - 444 , https://doi.org/10.1111/j.1365-2761.2004.00556.x https://doi.org/10.1111/j.1365-2761.2004.00556.x .

Li S J , Liu Y , Wang Y et al . 2019 . Lateral flow biosensor combined with loop-mediated isothermal amplification for simple, rapid, sensitive, and reliable detection of Brucella spp . Infection and Drug Resistance , 12 : 2343 - 2353 , https://doi.org/10.2147/idr.S211644 https://doi.org/10.2147/idr.S211644 .

Li Y M , Fan P H , Zhou S S et al . 2017 . Loop-mediated isothermal amplification (LAMP): a novel rapid detection platform for pathogens . Microbial Pathogenesis , 107 : 54 - 61 , https://doi.org/10.1016/j.micpath.2017.03.016 https://doi.org/10.1016/j.micpath.2017.03.016 .

Mori Y , Kanda H , Notomi T . 2013 . Loop-mediated isothermal amplification (LAMP): recent progress in research and development . Journal of Infection and Chemotherapy , 19 ( 3 ): 404 - 411 , https://doi.org/10.1007/s10156-013-0590-0 https://doi.org/10.1007/s10156-013-0590-0 .

Mori Y , Notomi T . 2020 . Loop-mediated isothermal amplification (LAMP): expansion of its practical application as a tool to achieve universal health coverage . Journal of Infection and Chemotherapy , 26 ( 1 ): 13 - 17 , https://doi.org/10.1016/j.jiac.2019.07.020 https://doi.org/10.1016/j.jiac.2019.07.020 .

Nagasawa K , Umino T , Grygier M J . 2007 . A checklist of the parasites of Ayu ( Plecoglossus altivelis altivelis ) (Salmoniformes: Plecoglossidae) in Japan ( 1912-2007 ). Journal of the Graduate School of Biosphere Science, Hiroshima University, 46 : 59 - 89 , https://doi.org/10.15027/30188. (in Japanese https://doi.org/10.15027/30188.(inJapanese )

Nair G , Rebolledo M , White A C Jr et al . 2015 . Detection of Entamoeba histolytica by recombinase polymerase amplification . The American Journal of Tropical Medicine and Hygiene , 93 ( 3 ): 591 - 595 , https://doi.org/10.4269/ajtmh.15-0276 https://doi.org/10.4269/ajtmh.15-0276 .

Notomi T , Mori Y , Tomita N et al . 2015 . Loop-mediated isothermal amplification (LAMP): principle, features, and future prospects . Journal of Microbiology , 53 ( 1 ): 1 - 5 , https://doi.org/10.1007/s12275-015-4656-9 https://doi.org/10.1007/s12275-015-4656-9 .

Nzelu C O , Kato H , Peters N C . 2019 . Loop-mediated isothermal amplification (LAMP): an advanced molecular point-of-care technique for the detection of Leishmania infection . PLoS Neglected Tropical Diseases , 13 ( 11 ): e 0007698 , https://doi.org/10.1371/journal.pntd.0007698 https://doi.org/10.1371/journal.pntd.0007698

Parida M , Sannarangaiah S , Dash P K et al . 2008 . Loop mediated isothermal amplification (LAMP): a new generation of innovative gene amplification technique; perspectives in clinical diagnosis of infectious diseases . Reviews in Medical Virology , 18 ( 6 ): 407 - 421 , https://doi.org/10.1002/rmv.593 https://doi.org/10.1002/rmv.593 .

Piepenburg O , Williams C H , Stemple D L et al . 2006 . DNA detection using recombination proteins . PLoS Biology , 4 ( 7 ): e 204 , https://doi.org/10.1371/journal.pbio.0040204 https://doi.org/10.1371/journal.pbio.0040204 .

Tu F , Yang X T , Xu S K et al . 2021 . Development of a fluorescent probe-based real-time reverse transcription recombinase-aided amplification assay for the rapid detection of classical swine fever virus . Transboundary and Emerging Diseases , 68 ( 4 ): 2017 - 2027 , https://doi.org/10.1111/tbed.13849 https://doi.org/10.1111/tbed.13849 .

Van De Peer Y , Ali A B , Meyer A . 2000 . Microsporidia: accumulating molecular evidence that a group of amitochondriate and suspectedly primitive eukaryotes are just curious fungi . Gene , 246 ( 1-2 ): 1 - 8 , https://doi.org/10.1016/s0378-1119(00)00063-9 https://doi.org/10.1016/s0378-1119(00)00063-9 .

Wittner M , Weiss L M . 1999 . The Microsporidia and Microsporidiosis . American Society for Microbiology, Washington . 572 p. https://do 10.1128/9781555818227 http://dx.doi.org/10.1128/9781555818227

Wong Y P , Othman S , Lau Y L et al . 2018 . Loop-mediated isothermal amplification (LAMP): a versatile technique for detection of micro-organisms . Journal of Applied Microbiology , 124 ( 3 ): 626 - 643 , https://doi.org/10.1111/jam.13647 https://doi.org/10.1111/jam.13647 .

Xiong Y F , Luo Y S , Li H et al . 2020 . Rapid visual detection of dengue virus by combining reverse transcription recombinase-aided amplification with lateral-flow dipstick assay . International Journal of Infectious Diseases , 95 : 406 - 412 , https://doi.org/10.1016/j.ijid.2020.03.075 https://doi.org/10.1016/j.ijid.2020.03.075 .

Zhou Q J , Chai F C , Chen J . 2018 . First record of Glugea plecoglossi (Takahashi Egusa , 1977 ), a microsporidian parasite of ayu (Plecoglossus altivelis altivelis Temminck Schlegel, 1846 ) in Mainland China. Journal of Fish Diseases , 41 ( 1 ): 165 - 169 , https://doi.org/10.1111/jfd.12674 https://doi.org/10.1111/jfd.12674 .

浏览量

Downloads

CSCD

文章被引用时，请邮件提醒。

Submit

工具集

关联资源

暂无数据

Construction and application of the RPA-LFD rapid detection method for Glugea plecoglossi

Construction and application of the RPA-LFD rapid detection method for Glugea plecoglossi

DOI：10.1007/s00343-024-4173-y

摘要

Abstract

关键词

Keywords

references