Blastocystis hominis has been regarded as an enigmatic parasite as many aspects of its basic biology remain uncertain. Many reproductive processes have been suggested for the organism; however, to date, only the binary fission has been proven. Plasmotomy is one of the modes of reproduction previously suggested to be seen in in vitro cultures. The present study provides trichrome and acridine orange staining evidence for the existence of nucleic acid suggestive of division of nucleus into multinucleate forms with the respective cytoplasm dividing giving rise to two or three progeny B. hominis. Transmission electron micrographs further confirmed that these daughter cells had respective surrounding surface coat, mitochondria, and vacuoles.
Genomic DNA from 16 Blastocystis hominis isolates comprising of eight asymptomatic isolates (A1-A8) and eight symptomatic isolates (S1-S8) was amplified by arbitrarily primed polymerase chain reaction (AP-PCR) using 38 arbitrary 10-mer primers. Six primers (A10, B5, C20, D1, F6, and F10) generated reproducible DNA fingerprints. AP-PCR amplification revealed similar DNA fingerprints among all symptomatic isolates (S1-S8) with common bands at 850 bp using primer A10, 920 bp using primer B5, and 1.3 kbp using primer D1. Isolates A1, A3, A4, A5, A6, and A7 showed similar DNA banding patterns and all asymptomatic isolates (A1-A8) shared a major band at 1 kbp using primer B5. Isolates A2 and A8 showed distinct DNA banding patterns that differed from the remainder of the isolates. The results of the phylogenetic analyses showed that all symptomatic isolates (S1-S8) formed a clade with >70% similarity among the isolates and which were clearly separate from asymptomatic isolates A1, A3, A4, A5, A6, and A7. Asymptomatic isolates A2 and A8 formed two distinct and separate clades. AP-PCR revealed higher genetic variability within the asymptomatic isolates than within the symptomatic isolates. The present study suggests that AP-PCR can be a valuable method for differentiating between isolates of B. hominis and our results support the hypothesis that our asymptomatic and symptomatic B. hominis isolates may represent two different strains/species with varying pathogenic potential.
The amoeboid form of Blastocystis hominis has been reported infrequently, and its morphological descriptions have yielded conflicting and confusing reports. In the present study, we used the amoeboid forms seen predominantly in symptomatic patients infected with Blastocystis to provide detailed descriptions on the fine surface structure and intracellular morphology. Scanning electron microscopy revealed the irregular shape of the amoeboid form, with an intercalated fibrillar structure and a highly convoluted surface with deep indentations and projected pseudopodia. Transmission electron microscopy showed the existence of two types of amoeboid forms of B. hominis in in vitro culture, one with a large central vacuole containing tiny electron-dense particles while the other contains multiple small vacuoles in the cytoplasm. A surface coat with varying thickness surrounded the amoeboid form, which also showed prominent, extended pseudopodia of varying shape. Irregularly shaped mitochondrion-like organelles with prominent cristae, lipid inclusions, and multiple vacuoles were frequently seen in close proximity with the pseudopodia. The characteristic nucleus with a crescentic band of electron-dense chromatin material was also seen.
Blastocystis hominis is one of the most common human parasites that inhabit the intestinal tract. Conflicting reports continue to exist regarding the existence and the functional role of the amoeboid forms in the life cycle of the parasite. The present study investigates the presence of these forms in 20 isolates obtained from ten symptomatic and asymptomatic patients respectively. A total of 10,000 parasite cells per ml from each isolate were inoculated into three culture tubes each containing 3 ml of Jones' medium supplemented with 10% horse serum, incubated at 37 degrees C. The contents were examined daily for 10 days. Irregular and polymorphic amoeboid forms with multiple extended pseudopodia were observed in all isolates from symptomatic patients, while none of the isolates from asymptomatic patients showed the presence of the amoeboid forms. The amoeboid forms were initially noted on day 2 and the percentages increased from 2% to 28%, with peak percentages from day 3 to day 6. Transmission electron microscopy revealed two types of amoeboid forms; one containing a large central vacuole completely filled with tiny electron-dense granules, and the other which revealed multiple small vacuoles within the central body. The cytoplasm contained strands of electron-dense granules resembling rough endoplasmatic reticulum, which is suggestive of active protein synthesis. The surface coat of the amoeboid form surrounding the parasite showed uneven thickness. Acridine orange stained the central body yellow and the periphery orange, indicating activity at the level of nucleic acids. The amoeboid form could either be an indicator of pathogenicity of B. hominis, or the form likely to contribute to pathogenicity and be responsible for the symptoms seen in patients.