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Objective <p>This study aimed to investigate the prevalence and subtype distribution of the genus <i>Blastocystis</i> using high-resolution melting curve analysis (HRM), a novel molecular technique. <i>Blastocystis</i> sp., a unicellular, anaerobic, and zoonotic parasite, is commonly found in the digestive tracts of humans and animals. Transmission occurs via the fecal-oral route, with prevalence rates ranging from 20% in developed countries to over 60% in developing regions. Although the pathogenicity of <i>Blastocystis</i> remains debated, it has been detected in both symptomatic and asymptomatic individuals. Traditionally, direct microscopy with wet mounts is used for the rapid identification of <i>Blastocystis</i> in stool samples. However, molecular diagnostics have become essential for the accurate detection and differentiation of <i>Blastocystis</i> subtypes</p> Materials and methods <p>We collected 730 stool samples from humans and domestic animals. Initial screening for <i>Blastocystis</i> sp. was performed using direct microscopy. Negative samples were cultured in a two-phase culture medium and re-examined after 2–3 days. HRM analysis was then employed to identify and differentiate <i>Blastocystis</i> subtypes using specific primers via real-time PCR.</p> Results <p>Six subtypes were identified, with ST7 (30%) being the most prevalent, followed by ST3 (28%), ST2 (16%), ST1 (14%), ST5 (6%), and ST14 (6%). In humans, ST3 was the most common subtype, also found in poultry and sheep, whereas ST7 was mainly detected in domestic animals. Notably, ST1-ST3 were identified in domesticated animals, indicating a pattern of cross-species transmission.</p> Conclusion <p>ST3 (28%) and ST7 (30%) were the most prevalent subtypes across all samples, with distinct distributions in human and animal hosts. The HRM technique demonstrated efficiency and cost-effectiveness, providing a rapid and accurate method for <i>Blastocystis</i> subtype identification in developing countries, which can expedite diagnostic responses and reduce the need for sequencing. This supports the potential for HRM to improve epidemiological surveillance and understanding of cross-species transmission.</p>

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Detection and subtyping of Blastocystis sp. in human and animal stool samples using high-resolution melting analysis

  • 期刊论文

  • 出版模式：

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  • 发表日期：

    2025年2月5日

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  • Ghodratollah Salehi Sangani,
  • Ahmad Hosseini-Safa,
  • Bibi Razieh Hosseini Farash,
  • Pooria Salehi Sangani,
  • Mehdi Zarean,
  • Elham Moghaddas,
  • Saeid Zoghdar Moghadam,
  • Lida Jarahi,
  • Ali Gholizadeh,
  • Fariba Berenji

摘要

Objective

This study aimed to investigate the prevalence and subtype distribution of the genus Blastocystis using high-resolution melting curve analysis (HRM), a novel molecular technique. Blastocystis sp., a unicellular, anaerobic, and zoonotic parasite, is commonly found in the digestive tracts of humans and animals. Transmission occurs via the fecal-oral route, with prevalence rates ranging from 20% in developed countries to over 60% in developing regions. Although the pathogenicity of Blastocystis remains debated, it has been detected in both symptomatic and asymptomatic individuals. Traditionally, direct microscopy with wet mounts is used for the rapid identification of Blastocystis in stool samples. However, molecular diagnostics have become essential for the accurate detection and differentiation of Blastocystis subtypes

Materials and methods

We collected 730 stool samples from humans and domestic animals. Initial screening for Blastocystis sp. was performed using direct microscopy. Negative samples were cultured in a two-phase culture medium and re-examined after 2–3 days. HRM analysis was then employed to identify and differentiate Blastocystis subtypes using specific primers via real-time PCR.

Results

Six subtypes were identified, with ST7 (30%) being the most prevalent, followed by ST3 (28%), ST2 (16%), ST1 (14%), ST5 (6%), and ST14 (6%). In humans, ST3 was the most common subtype, also found in poultry and sheep, whereas ST7 was mainly detected in domestic animals. Notably, ST1-ST3 were identified in domesticated animals, indicating a pattern of cross-species transmission.

Conclusion

ST3 (28%) and ST7 (30%) were the most prevalent subtypes across all samples, with distinct distributions in human and animal hosts. The HRM technique demonstrated efficiency and cost-effectiveness, providing a rapid and accurate method for Blastocystis subtype identification in developing countries, which can expedite diagnostic responses and reduce the need for sequencing. This supports the potential for HRM to improve epidemiological surveillance and understanding of cross-species transmission.