Application of modified JDP-DGGE-based molecular genotyping method to predict Acanthamoeba genotype and to analyse community diversity in aquatic environments

Tsui Kang Hsu, Jung Sheng Chen, Bing Mu Hsu, Yu Pin Chen, Tsai Hsueh Leu, Tung Yi Huang, Yu Wen Hsu, Shu Fen Wu

Research output: Contribution to journalArticle

1 Citation (Scopus)

Abstract

Acanthamoeba spp. are ubiquitous, opportunistic potential human pathogens, causing granulomatous amoebic encephalitis and keratitis. They are classified as protozoa, and they include at least 20 different genotypes (T1–T20) based on variation in the 18S rRNA gene. Acanthamoeba spp. are diverse in their production of toxins and in their ability to resist environmental factors. Therefore, it is necessary to develop a rapid genotyping method for Acanthamoeba spp. in aquatic environments. Although the denaturing gradient gel electrophoresis (DGGE) method for analysing microbial genotypes is potentially useful for rapid identification of aquatic environmental species, the technique has been compromised by artificial DGGE profiles in which many DNA fragments of identical sequences are segregated and displayed as different bands. The results indicate that PCR-DGGE genotyping with a GC clamp results in many segregated weaker bands of identical DNA sequences. In contrast, PCR-DGGE genotyping without a GC clamp displays genotype-dependent patterns in the major bands. Thus, DGGE without a GC clamp was performed to compare genotyping efficiency for Acanthamoeba in 21 water samples from rivers and reservoirs in Taiwan. Among them, four samples were found to demonstrate a banding pattern with more than one major band, and these band profiles of major bands were identical to those of positive controls. DNA cloning further confirmed that the sequences of the major bands were identical. In conclusion, more than two genotypes of Acanthamoeba in the four samples were identified by this method, suggesting that PCR-DGGE genotyping without a GC clamp is a useful approach for studying the diversity of Acanthamoeba communities. [Figure not available: see fulltext.].

Original languageEnglish
Pages (from-to)437-446
Number of pages10
JournalParasitology Research
Volume117
Issue number2
DOIs
Publication statusPublished - Feb 1 2018

Keywords

  • Acanthamoeba
  • Acanthamoeba communities
  • Genotype
  • Modified DGGE
  • Prediction

ASJC Scopus subject areas

  • Parasitology
  • veterinary(all)
  • Insect Science
  • Infectious Diseases

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