Journal of Microbiology and Biotechnology

The Korean Society for Microbiology and Biotechnology (한국미생물·생명공학회)
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The Use of the Internal Transcribed Spacer Region for Phylogenetic Analysis of the Microsporidian Parasite Enterocytozoon hepatopenaei Infecting Whiteleg Shrimp (Penaeus vannamei) and for the Development of a Nested PCR as Its Diagnostic Tool

  • Ju Hee Lee (Laboratory of Aquatic Biomedicine, College of Veterinary Medicine, Kyungpook National University) ;
  • Hye Jin Jeon (Laboratory of Aquatic Biomedicine, College of Veterinary Medicine, Kyungpook National University) ;
  • Sangsu Seo (Laboratory of Aquatic Biomedicine, College of Veterinary Medicine, Kyungpook National University) ;
  • Chorong Lee (Laboratory of Aquatic Biomedicine, College of Veterinary Medicine, Kyungpook National University) ;
  • Bumkeun Kim (Laboratory of Aquatic Biomedicine, College of Veterinary Medicine, Kyungpook National University) ;
  • Dong-Mi Kwak (Laboratory of Aquatic Biomedicine, College of Veterinary Medicine, Kyungpook National University) ;
  • Man Hee Rhee (Laboratory of Aquatic Biomedicine, College of Veterinary Medicine, Kyungpook National University) ;
  • Patharapol Piamsomboon (Department of Veterinary Medicine, Faculty of Veterinary Science, Chulalongkorn University) ;
  • Yani Lestari Nuraini (Fish Health and Environmental Laboratory, Brackishwater Aquaculture Development Center) ;
  • Chang Uook Je (Ministry of Agriculture, Food and Rural Affairs) ;
  • Seon Young Park (Division of Animal and Dairy Sciences, College of Agriculture and Life Science, Chungnam National University) ;
  • Ji Hyung Kim (Department of Food Science and Biotechnology, Gachon University) ;
  • Jee Eun Han (Laboratory of Aquatic Biomedicine, College of Veterinary Medicine, Kyungpook National University)
  • Received : 2024.01.10
  • Accepted : 2024.02.14
  • Published : 2024.05.28
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Abstract

The increasing economic losses associated with growth retardation caused by Enterocytozoon hepatopenaei (EHP), a microsporidian parasite infecting penaeid shrimp, require effective monitoring. The internal transcribed spacer (ITS)-1 region, the non-coding region of ribosomal clusters between 18S and 5.8S rRNA genes, is widely used in phylogenetic studies due to its high variability. In this study, the ITS-1 region sequence (~600-bp) of EHP was first identified, and primers for a polymerase chain reaction (PCR) assay targeting that sequence were designed. A newly developed nested-PCR method successfully detected the EHP in various shrimp (Penaeus vannamei and P. monodon) and related samples, including water and feces collected from Indonesia, Thailand, South Korea, India, and Malaysia. The primers did not cross-react with other hosts and pathogens, and this PCR assay is more sensitive than existing PCR detection methods targeting the small subunit ribosomal RNA (SSU rRNA) and spore wall protein (SWP) genes. Phylogenetic analysis based on the ITS-1 sequences indicated that the Indonesian strain was distinct (86.2% nucleotide sequence identity) from other strains collected from Thailand and South Korea, and also showed the internal diversity among Thailand (N = 7, divided into four branches) and South Korean (N = 5, divided into two branches) samples. The results revealed the ability of the ITS-1 region to determine the genetic diversity of EHP from different geographical origins.

Keywords

Acknowledgement

This research was supported by the Basic Science Research Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Education (NRF2019R1C1C1006212 and NRF-2022R1I1A3066435). The work was also supported by the Development of technology for biomaterialization of marine fisheries byproducts of the Korea Institute of Marine Science & Technology Promotion (KIMST) funded by the Ministry of Oceans and Fisheries (KIMST-20220128).

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