Small-subunit (SSU) rRNA gene sequences were obtained by PCR from 12 Blastocystis isolates from humans, rats, and reptiles for which elongation factor 1␣ (EF-1␣) gene sequences are already available. These new sequences were analyzed by the Bayesian method in a broad phylogeny including, for the first time, all Blastocystis sequences available in the databases. Phylogenetic trees identified seven well-resolved groups plus several discrete lineages that could represent newly defined clades. Comparative analysis of SSU rRNA-and EF-1␣-based trees obtained by maximum-likelihood methods from a restricted sampling (13 isolates) revealed overall agreement between the two phylogenies. In spite of their morphological similarity, sequence divergence among Blastocystis isolates reflected considerable genetic diversity that could be correlated with the existence of potentially >12 different species within the genus. Based on this analysis and previous PCR-based genotype classification data, six of these major groups might consist of Blastocystis isolates from both humans and other animal hosts, confirming the low host specificity of Blastocystis. Our results also strongly suggest the existence of numerous zoonotic isolates with frequent animal-to-human and human-to-animal transmissions and of a large potential reservoir in animals for infections in humans.

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Young (less than 8 weeks old) immunocompetent BALB/c mice became infected with Blastocystis hominis after inoculation of fecal cysts orally and of in vitro axenic-culture forms intracecally. This study confirmed that the fecal cyst was the form responsible for external transmission and that the mode of transmission was by the fecal-oral route. The infection was self-limiting and the infected BALB/c mice appeared normal except that some of them showed weight loss and lethargy. Both vacuolar and granular forms were found in the cecum, but only cyst forms were observed in the colon. Histological examination of the cecum and colon showed intense inflammatory-cell infiltration, edematous lamina propria, and mucosal sloughing. It is apparent that although B. hominis is not invasive, it is capable of causing pathogenesis in BALB/c mice.

Rather than using three layers, including an electrolyte, a working fuel cell is created that employs only one homogenous layer with mixed conductivity. The layer is a composite made from a mixture of metal oxide, Li0.15Ni0.45Zn0.4 oxide, and an ionic conductor; ion‐doped ceria. The single‐component layer has a total conductivity of 0.1–1 S cm−1 and exhibits both ionic and semiconducting properties. This homogenous one‐layer device has a power output of more than 600 mW cm−2 at 550 °C operating with H2 and air. Overall conversion is completed in a similar way to a traditional fuel cell, even though the device does not include the electrolyte layer critical for traditional fuel‐cell technologies using the three‐component anode–electrolyte–cathode structure.

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This paper elucidates the status of the different morphological forms of Blastocystis and reports the existence of thin- and thick-walled cysts in B. hominis on the basis of current experimental evidence. It is suggested that the thin-walled cysts are autoinfectious, leading to multiplication of the organism in the intestinal tract. The thick-walled cysts are responsible for external transmission via the faecal-oral route. A life cycle for B. hominis is postulated on the basis of these findings.

A fluorescence-based real-time 5 nuclease PCR capable of detecting all four human malaria parasites was developed to screen large numbers of samples during an outbreak to prevent further transmission of malaria. The effectiveness of antimalarial therapy for malaria patients can be monitored by determining the reduction of parasitemia by this method.Malaria is a major tropical infection causing up to 500 million clinical cases and 2.7 million deaths per year worldwide. For proper treatment of malaria patients, accurate and rapid diagnosis of malaria is essential. The microscopic examination of a blood smear is the "gold standard" for malaria diagnosis. The method is sensitive and specific but laborious and timeconsuming (6). Alternative methods, such as PCR and rapid antigen capture assays (ParaSight F test, OptiMAL rapid malaria test, and ICT Malaria P.f/P.v), have been developed. A PCR to detect Plasmodium falciparum and P. vivax was previously developed and evaluated (1,7,8,9). We have developed a method for malaria diagnosis that can be adapted for highthroughput rapid screening of hundreds of samples with a sensitivity and specificity comparable to those of the microscopic method. The method is a fluorescence-based real-time 5Ј nuclease PCR based on the TaqMan technology (Roche Molecular Diagnostics Systems) (3, 4), with primers and a probe from the small-subunit (SSU) rRNA gene.Malaria samples. Clinical blood samples were obtained from the Malaria Reference Center in the Department of Microbiology, National University of Singapore. Clinical samples of patients undergoing treatment were obtained from the National University Hospital, Singapore. The Gombak A strain of P. falciparum (5) was cultured by the method of Trager and Jensen (10). Two P. ovale (Po1 and Po2) and three P. malariae (Pm1, Pm2, and Pm3) samples were obtained.DNA extraction. DNA was extracted from 200 l of EDTAtreated blood samples with a QIAamp DNA blood minikit (Qiagen GmbH, Hilden, Germany) and stored at 4°C.Sequences of primers and probes. Primers (Mach 60 [5Ј-A CATGGCTATGACGGGTAACG-3Ј] and Mach 61 [5Ј-TGC CTTCCTTAGATGTGGTAGCTA-3Ј]) and a probe (Mach 62 [5Ј-TCAGGCTCCCTCTCCGGAATCGA-3Ј]) were designed to detect the SSU rRNA genes of all four human malarial species (accession numbers M19172 for P. falciparum, X13926 for P. vivax, M54897 for P. malariae, and L48987 for P. ovale). The probe was labeled with a reporter dye, FAM (5-carboxyfluorescein), and a quencher dye, TAMRA (N,N,N,N-tetramethyl-6-carboxyrhodamine) (Applied Biosystems, Foster City, Calif.). 5 nuclease PCR. Five microliters of DNA, 900 nM Mach 60, 300 nM Mach 61, 200 nM Mach 62, and 1ϫ TaqMan universal PCR master mix (Applied Biosystems) in a total reaction volume of 25 l were amplified on an iCycler (Bio-Rad Laboratories, Hercules, Calif.). The following conditions were used for PCR: an initial denaturation at 50°C for 2 min, 95°C for 10 min, and 45 cycles of 95°C for 15 s and 60°C for 1 min. The cycle threshold value (C T ) indicates the cycle where the fluorescence detect...