Limited molecular data exist on the prevalence and subtype distribution of Blastocystis spp., the most prevalent parasite in human and animal feces worldwide. A total of 44 different subtypes (STs) of Blastocystis are currently recognized based on the sequence of the small subunit ribosomal RNA (SSU-rRNA) gene. This is a molecular study of Blastocystis spp. in hospitalized patients with gastrointestinal symptoms in northern Spain. We analyzed 173 Blastocystis-positive patients with gastrointestinal symptoms by using nested PCR for molecular detection, subtype identification, phylogenetic analyses, and genetic diversity assessment. ST2 (34.1%) and ST3 (34.7%) predominated, followed by ST1 (15.6%) and ST4 (15.6%). Mixed infections with different subtypes were observed in some patients. Sequence analysis revealed for the first time in European humans the allele 88 (a variant of ST1). In other cases, alleles commonly found in animal samples were detected (allele 9 in ST2, allele 34 in ST3, and allele 42 in ST4). Phylogenetic analysis showed high variability in ST1 and ST2, suggesting a polyphyletic origin, while both ST3 and ST4 exhibited higher genetic homogeneity, indicating a possible monophyletic origin and recent transmission to humans. These data confirm Blastocystis spp. subtype diversity and may help in understanding the evolutionary processes and potential zoonotic transmission of this parasite.

Chlorine and solar disinfection are widely used disinfectants in water treatment. However, certain potential pathogens can resist these methods, posing a public health risk. One such case is Acanthamoeba, a resistant free-living amoeba that protects pathogens inside from disinfection, thus endangering the health of water users. This work is the first evaluation of the inactivation efficiency achieved by combining NaClO (Cl₂) and solar radiation (SR) against two Acanthamoeba strains from different sources (freshwater and pool water) and their endosymbiont bacteria (EB). Amoebae were exposed to different Cl₂ doses (0–500 mg/L), SR wavelength ranges (280–800 nm and 320–800 nm), used as gold standards, and their combinations. The EB exhibited resistance to conventional Cl₂ and SR treatments, requiring up to 20 times higher disinfectant doses than those needed to inactivate their protective Acanthamoeba. The pool strain and its EB demonstrated greater resistance to all treatments compared to the freshwater strain. Treatments with Cl₂ (5 mg/L)/SR_280–800nm completely inactivated both Acanthamoeba and EB of the freshwater strain, reducing up to 100 times the necessary Cl₂ doses, suggesting that chlorine photolysis is an attractive treatment for disinfecting freshwater and preventing waterborne diseases associated with Acanthamoebae and its EB.