Abstract
BackgroundBlastocystis sp. is one of the most common enteric parasites of humans and animals worldwide. It is well recognized that this ubiquitous protist displays a remarkable degree of genetic diversity in the SSU rRNA gene, which is currently the main gene used for defining Blastocystis subtypes. Yet, full-length reference sequences of this gene are available for only 16 subtypes of Blastocystis in part because of the technical difficulties associated with obtaining these sequences from complex samples.MethodsWe have developed a method using Oxford Nanopore MinION long-read sequencing and universal eukaryotic primers to produce full-length (> 1800 bp) SSU rRNA gene sequences for Blastocystis. Seven Blastocystis specimens representing five subtypes (ST1, ST4, ST10, ST11, and ST14) obtained both from cultures and feces were used for validation.ResultsWe demonstrate that this method can be used to produce highly accurate full-length sequences from both cultured and fecal DNA isolates. Full-length sequences were successfully obtained from all five subtypes including ST11 for which no full-length reference sequence currently exists and for an isolate that contained mixed ST10/ST14.ConclusionsThe suitability of the use of MinION long-read sequencing technology to successfully generate full-length Blastocystis SSU rRNA gene sequences was demonstrated. The ability to produce full-length SSU rRNA gene sequences is key in understanding the role of genetic diversity in important aspects of Blastocystis biology such as transmission, host specificity, and pathogenicity.Graphical
Highlights
Blastocystis sp. is one of the most common enteric parasites of humans and animals worldwide
Full-length sequences were successfully obtained from all five subtypes including ST11, for which no full-length reference sequence currently exists, and for an isolate that contained mixed ST10/ST14 (Table 1)
The ability to successfully generate full-length Blastocystis small subunit (SSU) ribosomal RNA (rRNA) gene sequences using MinION long-read sequencing technology from Blastocystis DNA extracted from culture and fecal samples was demonstrated
Summary
Blastocystis sp. is one of the most common enteric parasites of humans and animals worldwide. Is one of the most common enteric parasites of humans and animals worldwide. Is a common enteric protist parasite of humans and animals [1, 2]. It has a global distribution and is one of the most common human intestinal parasites in both developed and developing countries [3, 4]. Infection with Blastocystis in humans has been linked to gastrointestinal illnesses and/or urticaria [5, 6]. The pathogenicity of Blastocystis remains a topic of some controversy as asymptomatic infection is commonly reported [7, 8]. Blastocystis transmission occurs via the fecal-oral route. Many aspects related to the transmission and zoonotic potential of Blastocystis remain to be elucidated
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