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Background <p>Nested polymerase chain reaction (PCR) targeting the small subunit ribosomal ribonucleic acid (SSU rRNA, 18S rRNA) region is widely used to differentiate <i>Plasmodium</i> species. We identified a variant of the <i>Plasmodium malariae</i> SSU rRNA gene that suggests nested PCR may fail to detect <i>P. malariae</i> strains with unknown mutations.</p> Case presentation <p>A 56-year-old Japanese man developed a fever 2&#xa0;months after returning from a 2-month stay in Sierra Leone. Quartan malaria was suspected based on blood smear findings, and nested PCR confirmed <i>P. malariae</i> infection. However, the secondary PCR band obtained using <i>P. malariae</i>-specific primers was fainter than the primary PCR band amplified with universal primers—a reversal of the typical pattern. Sequence analysis revealed a four-base deletion in the SSU rRNA gene within the primer-binding site of the species-specific reverse primer used in the secondary PCR, suggesting that mutations in this region may partially impair amplification and hinder species identification. <i>Cytochrome b</i> gene sequencing confirmed 100% identity with <i>P. malariae</i>.</p> Conclusions <p>These findings underscore the need for continued molecular surveillance and sequence-based validation to ensure accurate diagnosis of <i>Plasmodium</i> infections, particularly in regions where genetic variants and zoonotic strains are emerging.</p>

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First report of a Plasmodium malariae SSU rRNA gene variant in Africa associated with reduced amplification by nested PCR

  • 期刊论文

  • 出版模式：

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  • 发表日期：

    2025年8月4日

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  • Maki Goto,
  • Kei Yamamoto,
  • Kanako Komaki-Yasuda,
  • Shigeyuki Kano,
  • Norio Ohmagari

摘要

Background

Nested polymerase chain reaction (PCR) targeting the small subunit ribosomal ribonucleic acid (SSU rRNA, 18S rRNA) region is widely used to differentiate Plasmodium species. We identified a variant of the Plasmodium malariae SSU rRNA gene that suggests nested PCR may fail to detect P. malariae strains with unknown mutations.

Case presentation

A 56-year-old Japanese man developed a fever 2 months after returning from a 2-month stay in Sierra Leone. Quartan malaria was suspected based on blood smear findings, and nested PCR confirmed P. malariae infection. However, the secondary PCR band obtained using P. malariae-specific primers was fainter than the primary PCR band amplified with universal primers—a reversal of the typical pattern. Sequence analysis revealed a four-base deletion in the SSU rRNA gene within the primer-binding site of the species-specific reverse primer used in the secondary PCR, suggesting that mutations in this region may partially impair amplification and hinder species identification. Cytochrome b gene sequencing confirmed 100% identity with P. malariae.

Conclusions

These findings underscore the need for continued molecular surveillance and sequence-based validation to ensure accurate diagnosis of Plasmodium infections, particularly in regions where genetic variants and zoonotic strains are emerging.