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Hisao Yoshikawa

Publications and source records attributed to Hisao Yoshikawa.

14 recordsLinked to original sources
PubMed2005

Infectivity of Blastocystis isolates from chickens, quails and geese in chickens.

The infectivity of six Blastocystis isolates obtained from two domestic chickens, two Japanese quails and two domestic geese, were examined in 1-week-old male chicks. All six isolates were able to infect the chicks via the intracecal inoculation of 1x10(6) cells of cultured organisms. Since the infected chicks discharged many cysts in their feces, the infectivity of the concentrated cysts in chicks was compared among three isolates from different bird species. The CK86-1 and QQ93-3 isolates, which were obtained from a chicken and a quail, respectively, were successfully infected in chicks by orally inoculating with 1x10(2)-1x10(6) cysts. On the other hand, the AC03-1 isolate from a goose required more cysts to infect the chicks, from 1x10(3) cysts to 1x10(6) cysts. In addition, when an uninfected normal chick was housed with five experimentally inoculated chicks with cysts of the QQ93-3 isolate, the normal chick became infected, indicating the fecal-oral transmission of the cyst form among the birds. These results show that the transmission of Blastocystis infection occurs easily between the same or different bird species. Therefore, the proposal of new Blastocystis species on the basis of different avian host species is problematic.

Animals↗
PubMed2004

Fecal-oral transmission of the cyst form of Blastocystis hominis in rats.

The infectivity of two Blastocystis hominis strains, RN94-9 and NIH:1295:1, was examined in 3-week-old SPF Wistar rats. The NIH:1295:1 strain, originally isolated from a guinea pig, was only able to infect rats via intracecal inoculation of the cultured organisms, while the RN94-9 strain, originally isolated from a laboratory rat, was able to infect rats by oral inoculation of the cultures due to the presence of a cystic form in the in vitro culture. Since many cysts were discharged in the feces of the infected rats, the infectivity of the concentrated cysts was compared between the two strains. Successful oral infection was observed in rats inoculated with 1 x 10(2)-1 x 10(6) cysts of the RN94-9 and NIH:1295:1 strains. The infectivity of the ten cysts varied in the three experiments of ten rats, being 20-100% and 30-100% in the RN94-9 and NIH:1295:1 strains, respectively. When an uninfected normal rat was housed with five experimentally inoculated rats, the normal rat became infected, demonstrating the fecal-oral transmission of the cyst form of this parasite. These results show that the Wistar rat is an ideal host for the propagation of strains RN94-9 and NIH:1295:1 of B. hominis, and demonstrate that the cyst form is the only transmissible form of this parasite.

Animals↗
PubMed2004

A survey of Blastocystis infection in anuran and urodele amphibians.

Blastocystis infection in amphibians was surveyed in three species of anuran and one species of urodele amphibians captured at two distinct locations in Japan. All three species of frogs were highly infected with Blastocystis, while 69 individual urodele newts, Cynopus pyrrhogaster, were negative for infection. Eleven Blastocystis isolates (47.8%) were recovered from 23 Rana nigromaculata leopard frogs. Twenty-three (92%) of 25 Rana catesbeiana bullfrogs and all (100%) of 24 Bufo japonicus japonicus toads were positive for Blastocystis. Two distinct populations of the toad and bullfrog showed a high prevalence (100 or 84.6%) of Blastocystis infection, while in two populations of the leopard frog only one population was positive for Blastocystis (84.6%). Three Blastocystis isolates from different species of the frogs were established. Since none of the three isolates could survive at 37 degrees C, a temperature tolerance assay was performed to assess the optimal growth temperature and to determine the range of non-lethal temperatures. During the exponential growth phase of 3- or 4-day cultures at 25 degrees C, three isolates were exposed to 4, 28, 31, or 34 degrees C for 3 days and then returned to 25 degrees C to monitor the cell growth. Based on the optimal growth temperatures and different ranges of temperature tolerance among the three new isolates from frogs and two known species, Blastocystis hominis and Blastocystis lapemi, it was established that the three isolates recovered from different species of frogs had different physiological features from B. hominis and B. lapemi.

Animals↗
PubMed2004

PCR-based identification of zoonotic isolates of Blastocystis from mammals and birds.

The genotype of Blastocystis isolated from humans and animals is highly polymorphic. Therefore, it is important to compare the genotypes of Blastocystis isolates from humans and animals to determine the zoonotic potential of animal isolates. PCR-based genotype classification using known sequence-tagged site (STS) primers allows identification of zoonotic isolates of animal origin. To this end, 51 isolates from monkeys, cattle, pigs, chickens, quails and pheasants were subjected to genotype analysis using seven kinds of STS primers. Out of the 51 isolates, 39 were identified as one of the known genotypes, four showed mixed genotypes, and eight were unknown genotypes as these were negative for all STS primers. When these results were combined with previous studies on 41 isolates from animals and compared with the diversity of genotypes of 102 human Blastocystis hominis isolates, 67.4 % (62/92) of isolates from mammals and birds were identical to human B. hominis genotypes. Since the unknown genotype of human origin had been placed into an additional clade in the small-subunit rRNA gene phylogeny, further molecular study on the eight isolates of unknown genotype from the present study will facilitate our understanding of their zoonotic potential.

Animals↗
PubMed2003

Polymerase chain reaction-based genotype classification among human Blastocystis hominis populations isolated from different countries.

Since the genotype of human Blastocystis hominis isolates is highly polymorphic, PCR-based genotype classification using known sequenced-tagged site (STS) primers would allow the identification or classification of different genotypes. Five populations of human B. hominis isolates obtained from Japan, Pakistan, Bangladesh, Germany, and Thailand were subjected to genotype analysis by using seven kinds of STS primers. Ninety-nine out of 102 isolates were identified as one of the known genotypes, while one isolate from Thailand showed two distinct genotypes and two isolates from Japan were negative with all the STS primers. The most dominant genotype among four populations, except for all four isolates from Thailand, was subtype 3 and it varied from 41.7% to 92.3%. The second most common genotype among four populations was either subtype 1 (7.7-25.0%) or subtype 4 (10.0-22.9%). Subtype 2, subtype 5, and/or subtype 7 were only rarely detected among the isolates from Japan and Germany, while subtype 6 was not detected. The phylogenetic position of the two isolates which were negative with all STS primers, was inferred from the small subunit rRNA (SSU rRNA) genes with the known sequence data of 20 Blastocystis isolates. Since the two isolates were positioned in an additional clade in the phylogenetic tree, this suggested they were a new genotype. These results demonstrated that PCR-based genotype classification is a powerful tool with which to analyse genotypes of Blastocystis isolates obtained from clinical samples. In addition, two groups of the isolates from 15 symptomatic and 11 asymptomatic patients in Bangladesh were compared with the PCR-based subtype classification. Since both groups were only classified into two distinct genotypes of subtype 1 or subtype 3 and no statistically significant difference was observed between the two groups, in this study it could not be shown that the specific genotype correlated with the pathogenic potential of B. hominis.

Animals↗
PubMed2003

Molecular characterization of Blastocystis isolates from primates.

Twelve Blastocystis isolates from primates were analyzed genetically by polymerase chain reaction (PCR) using diagnostic primers and PCR-restriction fragment length polymorphism (RFLP) of SSUrDNA. Two distinct genotypes, subtype 1 and a variant of subtype 1, were detected in two and six of the 12 isolates, respectively. The RFLP profiles of the isolates designated as subtype 1 were identical to the profile of ribodeme 1. The RFLP profiles of the six isolates designated as variants of subtype 1 were different from the profile of the variant of subtype 1 from a human reported previously. The other four isolates were not amplified with any diagnostic primers, but three of them showed the same RFLP profiles as ribodeme 6. This study was the first genomic analysis of Blastocystis isolates from primates, and showed the genetic similarity between the isolates from primates and the genotypes of Blastocystis hominis. However, it was unclear whether the isolates examined were zoonotic or not. Therefore, it is necessary to reveal the phylogenetic relationships between the isolates from primates and the multiple genotypes of B. hominis.

Animals↗
PubMed2003

Zoonotic genotypes of Blastocystis hominis detected in cattle and pigs by PCR with diagnostic primers and restriction fragment length polymorphism analysis of the small subunit ribosomal RNA gene.

Twenty two Blastocystis isolates from cattle and pigs were genotyped by PCR using diagnostic primers, and the homology among isolates was then confirmed by restriction fragment length polymorphism (RFLP) analysis of the small subunit ribosomal RNA gene. Two distinct genotypes, subtypes 1 and 3, were detected in one and two of the ten isolates from cattle, and three and one of the 12 isolates from pigs, respectively. The RFLP profiles of all isolates designated as subtype 1 and subtype 3 were identical to those of known ribodemes 1 and 2, respectively. We found that 31.8% (7/22) of isolates examined were zoonotic genotypes of Bastocystis hominis. However, 68.2% (15/22) of isolates were not amplified with any of the diagnostic primers, and the profiles of these isolates were different from any ribodemes of B. hominis reported previously. This study shows that Blastocystis organisms in cattle and pigs are a potential source of human infection.

Animals↗
PubMed2003

Molecular comparative studies among Blastocystis isolates obtained from humans and animals.

This study compared specific polymerase chain reaction (PCR) primers and phylogenetic tree analysis of restriction fragment length polymorphism using the small subunit ribosomal RNA gene in various Blastocystis populations. A phylogenetic tree was constructed using 12 restriction enzymes and a sample pool of 22 isolates, including 2 reference strains and Proteromonas lacertae as an outgroup. The analysis showed that the 22 isolates could be separated into 7 clusters. Four of the 7 clusters were mixed groups that comprised isolates from both humans and nonhuman hosts. The other 3 clusters contained isolates from humans or nonhuman hosts only. The phylogenetic analysis also showed that B. hominis isolates from geographical separated areas did not necessarily cluster in the genetically different groups. The results of genetic homology and phylogenetic tree analysis among Blastocystis isolates from humans and animals indicated that all isolates from animals appear to be B. hominis. Polymerase chain reaction amplifications using previously described and newly defined specific primers mirrored the clusters obtained by the phylogenetic tree analysis. Our results show that primer PCR can be used as a powerful tool for the typing of Blastocystis populations.

Animals↗
PubMed2002

Molecular characterization of Blastocystis isolates from birds by PCR with diagnostic primers and restriction fragment length polymorphism analysis of the small subunit ribosomal RNA gene.

Seven Blastocystis isolates from birds were genotyped by polymerase chain reaction (PCR) using diagnostic primers, and the homology among isolates was then confirmed by restriction fragment length polymorphism (RFLP) analysis of the small subunit ribosomal RNA gene. A zoonotic genotype, subtype 4, was detected in three isolates by PCR, but the RFLP profiles of these isolates differed from those of the subtype 4 reported previously from a human. None of the other four isolates were amplified with any subtype-specific primer sets, and the profiles of these isolates were different from any ribodemes of Blastocystis hominis reported previously. In the present study, the seven isolates from birds were classified into three genetically distinct genotypes. Therefore, birds appear to be infected with multiple genotypes. It is unclear whether the isolates examined in the present study are zoonotic or not. Therefore, it is important to determine the phylogenetic relationships between unknown genotypes from birds and the multiple genotypes of B. hominis.

Animals↗
PubMed2002

A survey of Blastocystis sp. in livestock, pets, and zoo animals in Japan.

The prevalence of Blastocystis sp. was examined in fecal samples collected from cattle, pigs, dogs, and a variety of zoo animals (primates, carnivores, herbivores, pheasants, and ducks) by direct observation of fresh fecal suspensions or cultured materials, using light microscopy. The cattle and pigs were randomly sampled from 11 and 12 commercial farms, respectively, located in the western region of Japan. The dog material used in this study was obtained from pets housed in an animal shelter in the city of Osaka. Zoo animals were chosen based on housing conditions that minimized the possibility of intra-zoo transmission of the organism. The prevalence rate among the groups varied greatly. A high prevalence of infection was observed in the farm animal group, ranging from 95% (58/61) in pigs to 71% (39/55) in cattle, whereas the dog fecal samples were completely free of the organism. Prevalence of the organism in the zoo animal were 85% (29/34) in primates, 80% (8/10) in pheasants, 56% (9/16) in ducks, and 0% (0/58) in various carnivores and herbivores. Among the zoo animals infected with Blastocystis, eight species of primates, eight species of pheasants, and four species of ducks were confirmed as new hosts. Since Blastocystis organisms isolated from various animals were morphologically indistinguishable from Blastocystis hominis by light microscopy, further genomic studies are required for analysis of the zoonotic potential or etiological significance of these isolates.

Animals↗
PubMedDate not supplied

Phylogenetic position of Blastocystis hominis and of stramenopiles inferred from multiple molecular sequence data.

Blastocystis hominis, a parasite of the human intestine, has recently been positioned within stramenopiles by the small subunit rRNA phylogeny. To further confirm its phylogenetic position using multiple molecular sequence data, we determined the nucleotide sequences putatively encoding small subunit ribosomal RNA, cytosolic-type 70-kDa heat shock protein, translation elongation factor 2, and the non-catalytic 'B' subunit of vacuolar ATPase of B. hominis (HE87-1 strain). Moreover, we determined the translation elongation factor 2 sequence of an apicomplexan parasite, Plasmodium falciparum, that belongs to alveolates. The maximum likelihood analyses of small subunit rRNA and cytosolic-type 70-kDa heat shock protein clearly demonstrated that B. hominis (HE87-1 strain) is positioned within stramenopiles, being congruent with the previous small subunit rRNA analysis, including the sequences of B. hominis (Nand strain) and a Blastocystis isolate from guinea pig. Although no clear resolution among major eukaryotic groups was obtained by the individual phylogenies based on the four molecules analyzed here, a combined analysis of various molecules, including these, clearly indicated that Blastocystis/stramenopiles are the closest relatives of alveolates.

Animals↗
PubMedDate not supplied

Freeze-fracture and cytochemical studies on the in vitro cyst form of reptilian Blastocystis pythoni.

After cultured cysts are osmotically shocked by treating with distilled water, there is an exponential increase in the cyst form of Blastocystis pythoni; this was demonstrated by an immunofluorescence antibody assay against the culture organisms. In 11-day-old cultures of B. pythoni, 68.8% of the organisms (= 2.2 x 10(8) cysts/ml) were in the cyst form. Examination of thin sections of cysts revealed many similarities to the cyst forms of Blastocystis obtained from fecal samples in previous investigations. Freeze-fracture images of the plasma membrane of non-cyst cells also revealed a similar distribution of the intramembrane particles (IMP) when compared to non-cysts of B. hominis, while the plasma membrane of the cyst form showed practically no IMP. The size and morphology of particle-rich small depressions and smooth small protrusions observed on the P face and E face of non-cyst cells, respectively, were similar to endocytic sites reported for B. hominis. In the present study glycogen was cytochemically demonstrated at the ultrastructural level by an alkaline bismuth staining method in both cyst and non-cyst cells.

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