Hideto Kino

Sphingolipids of a cestode Metroliasthes coturnix

Sphingolipids of Metroliasthes coturnix were studied. The cestode contained no detectable amounts of sphingomyelin. The major glycosphingolipids found were monogalactosylceramide, galactosyl-alpha-1-4-galactosylceramide, galactosyl-beta-1-6-galactosyl-beta-1-6-galactosylceramide and galactosyl-beta-1-6-galactosyl-beta-1-6-galactosyl-beta-1-6-galactosylce ramide. Their ceramides were mostly composed of C18-20 sphinganine or 4-D-hydroxysphinganine, which is N-acylated by extraordinarily long normal and 2-hydroxy acids such as C26 acid (range C16 to C30) except tetragalactosylceramide. The structure of glycosphingolipids was confirmed after their chromatographic separation into each molecular species using a novel analytical device, fast atom bombardment-mass spectrometry linked with reversed-phase high-performance liquid chromatography.

A mass occurrence of human infection with Diplogonoporus grandis (Cestoda: Diphyllobothriidae) in Shizuoka Prefecture, central Japan.

Forty-six cases of human infection with Diplogonoporus grandis were found in Shizuoka Prefecture on the Pacific coast of Central Japan in 1996. The cases were predominantly elderly male patients over 50 years of age. Although all cases were reported from May to September of the year, most of them were diagnosed in June and July. We suspected that the transmission was due to the consumption of raw juvenile Japanese anchovy (Engraulis japonicus), which are seasonally caught in the spring months off the Pacific coast of the Prefecture. In almost a hundred years after its discovery in 1894, there had been more than 180 cases of human diplogonoporiasis recorded in Japan. The high incidence within a relatively short time frame of our investigation is regarded unusual even in this country.

Angiostrongylus cantonensis: Paralysis due to avermectin B1a and ivermectin

Paralysis due to avermectin B1a and ivermectin of Angiostrongylus cantonensis was compared to that of phenylephrine (an alpha-adrenergic agonist) and strychnine (a cholinergic inhibitor). The paralyzing action of ivermectin (2.5 X 10(-9) g/ml) was inhibited by the single, simultaneous addition of picrotoxin (3 X 10(-5) M), whereas the effect of the drug (2.5 X 10(-7) g/ml) was reversed only when picrotoxin was given with cholinergic spasmogens such as pyrantel and eserine. Bicuculline (3 X 10(-5) M) had a similar antagonistic effect for picrotoxin, but bicuculline was less effective. The paralyzing action of avermectin B1a (3.6 X 10(-14) M, 3.0 X 10(-14) g/ml) was antagonized only when picrotoxin was given with cholinergic spasmogens such as pyrantel, eserine, and N-methylcytisine (N-MC), or alpha-adrenergic antagonists such as phentolamine and dibenamine. On the other hand, the paralyzing action of strychnine (3 X 10(-6) M) or phenylephrine (3 X 10(-5) M) was relatively uninfluenced by picrotoxin, but was antagonized by pyrantel and N-MC or dibenamine. These results suggest that a gabergic mechanism is involved in the paralyzing action of ivermectin, as well as avermectin B1a, in A. cantonensis.

Studies on chemotherapy of parasitic helminths: Effects of avermectin Bla on Angiostrongylus Cantonensis in rats

Abstract The in vivo effects of avermectin Bla against Angiostrongylus cantonensis in rats were examined. Two distinguishable effects of this drug were suggested; a paralyzing effect against adult worms and a direct vermicidal effect against larval worms. When the drug was given intraperitoneally at 0·01 or 0·1 mg/kg at 6 weeks after infection, a sustained paralysis of worms by this drug was suggested from the following aspects; the change of the first stage larval count in feces, histological observations of lung tissues of the host and the reproductive system of female worms, and the motility of recovered worms in vitro . However, no direct vermicidal effect was observed in this treatment, because there was no significant difference in the recovery rate of worms from that of control rats. On the other hand, when the drug was given orally at a dose of 1·0 mg/kg at 3 days after infection (larval stage treatment), a significant reduction in the recovery rate of worms was observed compared to those in the adult stage treatment (drug administration at 7 weeks after infection) and control groups.

Resistance of a rodent malaria parasite to a thymidylate synthase inhibitor induces an apoptotic parasite death and imposes a huge cost of fitness.

Background The greatest impediment to effective malaria control is drug resistance in Plasmodium falciparum, and thus understanding how resistance impacts on the parasites fitness and pathogenicity may aid in malaria control strategy. Methodology/Principal Findings To generate resistance, P. berghei NK65 was subjected to 5-fluoroorotate (FOA, an inhibitor of thymidylate synthase, TS) pressure in mice. After 15 generations of drug pressure, the 2% DT (the delay time for proliferation of parasites to 2% parasitaemia, relative to untreated wild-type controls) reduced from 8 days to 4, equalling the controls. Drug sensitivity studies confirmed that FOA-resistance was stable. During serial passaging in the absence of drug, resistant parasite maintained low growth rates (parasitaemia, 15.5%±2.9, 7 dpi) relative to the wild-type (45.6%±8.4), translating into resistance cost of fitness of 66.0%. The resistant parasite showed an apoptosis-like death, as confirmed by light and transmission electron microscopy and corroborated by oligonucleosomal DNA fragmentation. Conclusions/Significance The resistant parasite was less fit than the wild-type, which implies that in the absence of drug pressure in the field, the wild-type alleles may expand and allow drugs withdrawn due to resistance to be reintroduced. FOA resistance led to depleted dTTP pools, causing thymineless parasite death via apoptosis. This supports the tenet that unicellular eukaryotes, like metazoans, also undergo apoptosis. This is the first report where resistance to a chemical stimulus and not the stimulus itself is shown to induce apoptosis in a unicellular parasite. This finding is relevant in cancer therapy, since thymineless cell death induced by resistance to TS-inhibitors can further be optimized via inhibition of pyrimidine salvage enzymes, thus providing a synergistic impact. We conclude that since apoptosis is a process that can be pharmacologically modulated, the parasites apoptotic machinery may be exploited as a novel drug target in malaria and other protozoan diseases of medical importance.

Chromosomal mapping of host resistance loci to Trichinella spiralis nematode infection in rats.

The differences in host response among strains of rats to intestinal nematode parasite Trichinella spiralis infection could provide a powerful benefit for further elucidation of molecular interactions between the host and the parasite. Using several strains of rats, we previously observed that DA strain is a strong responder and F344 strain is a weak responder with respect to expulsion of the adult worm. To identify the host resistance loci, quantitative trait loci (QTLs) analysis in F2 population from crosses between DA and F344 strains was performed. One significant QTL (designated as Tspe) was mapped to the middle region of chromosome 9. In addition, the effect of DA allele at Tspe locus could act recessively and lead to the rejection of more adult worms from the gut. The results from the present study provide more insights on host–parasite interactions, which may be useful in facilitating the development of novel approaches for treatment and control of intestinal parasites in human and domestic livestock.

Effects of Avermectin Bla on the motility of various parasitic helminths

Using an isotonic transducer recently devised in Japan, it was observed that Avermectin Bla, a new anthelmintic, caused paralyzing effects onAngiostrongylus cantonensis andMetastrongylus elongatus at concentrations of more than 3.6×10−18 M.

A molecular phylogeny of Asian species of the genus Metagonimus (Digenea)—small intestinal flukes—based on representative Japanese populations

Metagonimus Katsurada, 1912 is a genus of small intestinal parasites. The genus comprises eight species, primarily from far-eastern Asia, with two exceptions reported from Europe. Metagonimus yokogawai, the most widespread species, is the main agent responsible for the intestinal disease, metagonimiasis, in Japan and some other East Asian countries. On the basis of the ratio of the size of the ventral and oral suckers, Metagonimus has traditionally been morphologically divided into two groups; however, the genus has not been extensively studied using molecular data. To reveal phylogenetic relationships within Metagonimus based on molecular data, we analyzed six of the seven species present in Asia using samples collected in central Japan. Maximum likelihood and Bayesian analyses of a combined 28S ribosomal DNA (rDNA), internal transcribed spacer 2 (ITS2), and mitochondrial cox1 gene sequence dataset separated the six species into two well-supported clades. One clade comprised M. yokogawai, M. takahashii, M. miyatai, and M. hakubaensis, whereas the other consisted of M. otsurui and M. katsuradai. Genetic distances calculated from 28S rDNA and ITS2 nucleotide sequences and a comparison of the predicted amino acid sequences of cox1 gene suggested that M. otsurui and M. katsuradai may have diverged recently. None of the four main morphological characters used to delimit species of Metagonimus (i.e., sucker ratio, positions of the uterus and testes, and distribution of vitelline follicles) was consistent with the distribution of species in the molecular tree.

Effects of mebendazole onAngiostrongylus costaricensis in mice, with special reference to the timing of treatment

Mebendazole was given to mice infected withAngiostrongyls costaricensis at a single dose of 5 mg/kg at 6, 11, 16 or 21 days post-infection (p.i.) and in five successive doses at 5 mg/kg daily at 6, 11 or 16 days p.i. The effects were comparatively assessed by examining various parameters in host mice and worms. As a whole, the effects of mebendazole were caused more conspicuously by five successive treatments than by a single treatment. In both treatment modalities, the effects were more remarkable in earlier treatments, and nearly complete effects were caused by five successive treatments before 15 days p.i. These results suggest that the inhibition of egg formation and/or oviposition will inhibit the pathological changes caused in the disease byA. costaricensis, especially before the onset of the changes.

Studies on chemotherapy of parasitic helminths (XVIII). Mechanism of spastically paralyzing action of pyrantel in Angiostrongylus cantonensis

Pyrantel tartrate caused spastic paralysis through stimulating nicotinic cholinoceptors inAngiostrongylus cantonensis.