Efficacy of thiabendazole, mebendazole, levamisole and ivermectin against gullet worm, Gongylonema pulchrum: in vitro and in vivo studies

In vitro and in vivo studies were conducted to evaluate the effects of thiabendazole, mebendazole, levamisole and ivermectin against Gongylonema pulchrum. For in vitro assays, third-stage larvae (L3) incubated with the drugs were administered orally to mice and the ability of larvae to invade the gastric mucosa of the animals was examined. After incubation, only those larvae treated with high concentrations of levamisole (1 and 10 microg/ml) were tightly coiled with intestines exhibiting morphological abnormalities. Good dose-response data for the drugs tested was observed at the time of worm recovery from mice, with no worms recovered at the two highest concentrations of levamisole. In vivo efficacy of the drugs against adult worms was evaluated in six groups of three rabbits, each of which was infected with 30 L3 of G. pulchrum and treated with thiabendazole at 100 mg/kg for 3 days, mebendazole at 70 mg/kg for 3 days, levamisole as a single dose of 8 mg/kg, and subcutaneously injected ivermectin as a single dose of 0.2 mg/kg or vehicles of the drugs (control) at 4 months post-infection. Necropsy 14 days after treatment revealed that levamisole, mebendazole and ivermectin reduced worm burdens by 63.2%, 22.8% and 25.8%, respectively, with no reductions in worms observed with thiabendazole. The surviving worms were principally found in the esophagus with the remainder distributed among the buccal mucosa, the tongue, and/or pharyngeal mucosa in all groups. A number of morphologically abnormal eggs were observed within the uterus and ovijector in female worms recovered from the thiabendazole-treated group. These findings suggest that levamisole exhibits in vivo efficacy against G. pulchrum infection and that the larval invasion tests using mice could be used to screen for anthelmintic susceptibility of nematodes.     

In vitro evaluation of the growth inhibitory activities of 15 drugs against Babesia gibsoni (Aomori strain)

The in vitro growth inhibitory activities of 15 drugs against Babesia gibsoni were evaluated following establishment of a continuous culture isolate (Aomori isolate). The culture was successfully continued in an RPMI-1640 medium supplemented with 20% normal canine serum or fetal bovine serum in a humidified atmosphere containing 5% CO(2) and 5% O(2) at 37 degrees C. We used this isolate to evaluate the growth inhibitory effect of naphthoquinone (atovaquone), aromatic diamidine (diminazene and pentamidine), artemisinin compounds (artesunate and dihydroartemisinin), an iron chelator (deferoxamine), quinoline-containing compounds (quinine and chloroquine), macrolide antibiotics (azithromycin), lyncomycin antibiotics (clindamycin), tetracycline antibiotics (doxycycline and minocycline), imidazole antifungals (clotrimazole and ketoconazole), and a nitroimidazole antiprotozoal (metronidazole). Atovaquone and aromatic diamidine showed the highest activity; they were followed by artesunate compounds with nanomole levels of IC(50). Metronidazole did not exhibit activity against the parasite. Other drugs exhibited intermediate in vitro activities with micromole levels of IC(50). This is the first report to screen drug activities against B. gibsoni in vitro. The results of our study may support further in vitro drug evaluation for the establishment of therapeutic strategies against canine B. gibsoni infections.     

Retrospective diagnosis of feline GM2 gangliosidosis variant 0 (Sandhoff-like disease) in Japan: possible spread of the mutant allele in the Japanese domestic cat population

GM2 gangliosidosis variant 0 (human Sandhoff disease) is a lysosomal storage disease caused by simultaneous deficiencies of acid beta-hexosaminidase (Hex) A and Hex B due to an abnormality of beta-subunit, a common component in these enzyme molecules, which is coded by the HEXB gene. In the present study, a retrospective diagnosis was performed in 2 previous suspected cases of feline Sandhoff-like disease using a DNA test to detect the causative mutation identified previously in 4 cats in 2 other families of Japanese domestic cats. Enzymic analysis was also performed using stored leukocytes and plasma collected from the subject families in order to investigate the usefulness of enzymic diagnosis and genotyping of carriers. The DNA test suggested that the 2 cases were homozygous recessive for the mutation. Consequently, 6 cats homozygous for the same mutation have been found in 4 separate locations of Japan, suggesting that this mutant allele may be spread widely in the Japanese domestic cat populations. In enzymic analysis, Hex A and Hex B activities in leukocytes and plasma measured using 4-methylumbelliferyl N-acetyl-beta-D-glucosaminide as a substrate were negligible in affected cats, compared with those in normal and carrier cats. However, there was a wide overlap in enzyme activity between normal and carrier cats. Therefore, it was concluded that enzymic analysis is useful for diagnosis of affected cats, but is not acceptable for genotyping of carriers.     

New lactic acid bacteria for skin health via oral intake of heat‐killed or live cells

Lactic acid bacteria play an essential role in the food industry in the manufacture of many fermented products (cheese, yogurt, fermented vegetables, etc.). Application of these organisms is now being extended to the area of health improvement, as their probiotic activities become known. Probiotics are defined as viable microorganisms that exert a beneficial effect on the health of the host when they are ingested in sufficient quantity. Lactic acid bacteria and bifidobacteria isolated from the human intestine are the most common probiotics used for human consumption. The development of new probiotics with new beneficial effects is eagerly awaited in the food industry. This review introduces Lactococcus, which are one of the genera of lactic acid bacteria and are mainly isolated from dairy products and fermented vegetables, as new probiotics, focusing especially on Lactococcus lactis H61, which improves skin status in Japanese women with oral intake of heat‐killed or live cells. The deduced mechanisms associated with the beneficial effects of strain H61 are also discussed.     

Production of androgenetic amago salmon<Emphasis Type="Italic"> Oncorhynchus masou ishikawae</Emphasis> with dispermy fertilization

With the aim of improving artificial androgenesis in teleost fishes, we tested two methods for producing androgenetic diploids of amago salmon (Oncorhynchus masou ishikawae), namely, fertilization of gamma-ray irradiated eggs with fused spermatozoa (sperm-fusion method) and the fertilization of irradiated eggs with untreated sperm followed by the blocking of cell division (mitosis-inhibition method). Our results showed that the optimal condition for sperm fusion was to treat the sperm with 50% polyethylene glycol (molecular weight 7500) for 100 s. The efficiency of the two methods of androgenesis was compared in terms of fertilization rate, hatching rate, and larval survival after hatching. The rate of fertilization was lower with the sperm-fusion method than with the mitosis-inhibition method, but the reverse was true for the hatching rate. The survival rate of hatched larvae was the same with the two methods. Androgenesis was confirmed with a recessive albino color marker, and all viable offspring were found to be heterozygous based on analysis of the microsatellite markers. Our results suggest that androgenesis with the sperm-fusion method is a promising approach with potential applications in both aquaculture breeding programs and the preservation of endangered freshwater fishes.     

Biochemical comparison of lectins among three different color strains of the red alga Kappaphycus?alvarezii

The red alga Kappaphycus alvarezii is economically important as an edible species and as a source of carrageenan, and has been extensively cultivated in many tropical countries. For this species, different color strains, which differ from each another in growth rate and carrageenan content, have been reported for decades. In this study, lectins from brown, red, and green strains of K. alvarezii cultivated in Vietnam were isolated and characterized for evaluation of their biochemical properties and contents. The results showed that each color strain contained in common the three lectins, named KAA-1, KAA-2, and KAA-3, which shared the hapten-inhibition profile of hemagglutination, 20 N-terminal amino acid sequence, and equivalent molecular mass within a range of 28,016 ± 1.2 to 28,021 ± 1.8 Da, but differed in their yields, with the highest yield of KAA-2. These properties of the three isolectins were also comparable among the three different color strains. However, the sum of the yields of the three isolectins decreased in the order: red (21.4 mg) to green (15.9 mg) to brown strains (15.1 mg), from 500 g fresh alga. Thus, this algal species can be a good source of useful lectins, irrespective of color strain.     

Mechanism of the rice hull‐induced germination of Monochoria vaginalis seeds in darkness

The seeds of Monochoria vaginalis require light to germinate. However, they are able to germinate in darkness in the presence of rice hulls. The mechanism of this phenomenon was investigated in this study, in which only the unsterilized seeds of Monochoria were induced to germinate by the presence of a rice hull extract. The rice hull extract was able to be replaced by either a mixture of amino acids and phosphate or a yeast extract. The culture media in which germination was observed always showed turbidity, indicating the propagation of bacteria, which were identified as Bacillus sp., Pedobacter sp., Pantoea sp. and Sphingomonas sp. Treatment with the individual bacterial species or combinations of these bacteria induced Monochoria seed germination when added to the media containing the rice hull extract. It was concluded that the rice hull extract accelerates the propagation of bacteria, which in turn digest the seed coat of Monochoria, facilitating its germination.     

Psycho-educational Horseback Riding to Facilitate Communication Ability of Children with Pervasive Developmental Disorders

In this study, we applied a novel psycho-educational horseback riding (PEHR) program to the treatment of four Japanese children with pervasive developmental disorders (PDD) in order to facilitate the acquisition of verbal and nonverbal communication skills. The behavioral changes in each child were evaluated using a psychological and behavioral scale. The scale for evaluating the effect of Human-Equips-Interaction on Mental activity (HEIM scale) was designed to assess the behavioral improvement of children based on the following 10 items: Human relationships, Imitation, Emotional expression, Sudden physical movement, Fixative behavior, Adaptation to change, Visual response, Fear or nervousness, and Verbal and nonverbal communication. After taking part in the PEHR program for several months, all subjects showed remarkably improved HEIM scores and marked improvements were observed in eye contact with others (instructors, side walkers, and leaders) in the riding area. A statistical difference was found in items 1, 2, 3, 6, 7, 8, and 9. However, no statistical difference was found in items 4, 5, and 10. As the program progressed, the children showed enhanced verbal and nonverbal communication skills, and became more expressive in their emotional and empathetic interaction with their parents. These observations suggest that the normal functioning of pleasurable emotions and empathy may facilitate further improvements in joint attention, imitation and empathy, and may result in successful verbal expression by PDD children. Therefore, horseback riding can play a very important role in the psycho-educational support required for the communication ability of PDD children.     

An avirulence gene to rice cultivar K60 is located on the 1.6-Mb chromosome in <Emphasis Type="Italic">Magnaporthe oryzae</Emphasis> isolate 84R-62B

A Japanese differential rice cultivar K60 was tested with 114 F₁ cultures of Magnaporthe oryzae from a cross between isolates 84R-62B and Y93-245c-2. Segregation patterns of avirulence and virulence in the progeny suggested that avirulence on cv. K60 was controlled by a single gene derived from 84R-62B and tentatively named AvrK60. In the F₁ population, AvrK60 cosegregated with avirulence gene AvrPik on a small 1.6-Mb chromosome of 84R-62B and with the 1.6-Mb chromosome itself. Therefore, we suggest that, along with AvrPik, AvrK60 is also located on the 1.6-Mb chromosome of 84R-62B.     

Effects of cooling rate on post-thaw motility and fertility of Japanese pearl oyster <Emphasis Type="Italic">Pinctada fucata martensii</Emphasis> spermatozoa

The post-thaw motility and fertility of Japanese pearl oyster sperm show large variances, even among sperm samples obtained from the same individuals. This study aimed to clarify the factors that cause such differences. Spermatozoa were diluted 50 times with diluent comprising 10 % methanol, 18 % fetal bovine serum, and 72 % seawater, and dispensed into 0.25 ml straws. A total of 59 straws were cooled, one by one, at 11 different heights from the surface of liquid nitrogen (LN) to −50 °C, and then immediately immersed in LN. After thawing the straws, the relationships between the cooling rate and the post-thaw motility and post-thaw fertility of the spermatozoa were examined. Both the post-thaw motility and the post-thaw fertility showed a sharp peak when the straws were cooled at around −20 °C/min. There was a strong correlation between post-thaw motility and fertility (P < 0.001). There was a large difference in the cooling rates and the post-thaw motilities and fertilities of the spermatozoa, even between straws cooled at the same height. These results indicate that the optimum range for the cooling rate of oyster spermatozoa is quite narrow, and the method of cooling straws at a fixed distance from the LN surface is unsuitable for the cryopreservation of Japanese pearl oyster spermatozoa.