Pathological findings of Japanese Black Cattle living in the restricted area of the Fukushima Daiichi Nuclear Power Plant accident, 2013–2016

Fifty‐one Japanese black cattle from four farms in the evacuation zone of the Fukushima nuclear accident were examined pathologically during the period, 2013–2016. We found no evidence of a radiation effect on pathological findings in any of these autopsy cases, although nine cases (3.7%) of enzootic bovine leukosis (EBL) and three cases (1.2%) of goiter were diagnosed. Estimated integrating dose of external exposure in EBL cases ranged from a maximum of 1200 mSv to a minimum of 72 mSv. Clinically, five cases showed wobble, dysstasia or paralysis. Exophthalmos was observed in three cases. Macroscopically, enlarged lymph nodes, multiple irregular masses of intra‐abdominal and intrapelvic adipose tissue, diffuse thickening of the abomasal submucosa with ulceration, and numerous white nodules of myocardium were observed. Histologically, neoplastic lymphoid cells were extensively proliferated and infiltrated in the lesions. Lymphoid tumor cells showed mature or large lymphoblastic appearance. Immunohistochemically, tumor cells were positive for BLA‐36, CD20 and CD5, and negative for CD3. Three cases showed diffuse enlargement of the thyroid gland with no mass or nodule lesion. Histologically, diffuse proliferation of follicular epithelium was observed with preservation of normal structures. There were no malignant findings such as cellular atypia or invasion to capsule.     

Site‐directed mutagenesis of the cytochrome b gene and development of diagnostic methods for identifying QoI resistance of rice blast fungus

BACKGROUND: It is possible that a single nucleotide polymorphism (SNP) (G143A mutation) in the cytochrome b gene could confer resistance to quinone outside inhibiting (QoI) fungicides (strobilurins) in rice blast fungus because this mutation caused a high level of resistance to fungicides such as azoxystrobin in Pyricularia grisea Sacc. and other fungal plant pathogens. The aim of this study was to survey Magnaporthe oryzae B Couch sp. nov. isolates in Japan for resistance to QoIs, and to try to develop molecular detection methods for QoI resistance. RESULTS: A survey on the QoI resistance among M. oryzae isolates from rice was conducted in Japan. A total of 813 single‐spore isolates of M. oryzae were tested for their sensitivity to azoxystrobin using a mycelial growth test on PDA. QoI fungicide resistance was not found among these isolates. The introduction of G143A mutation into a plasmid containing the cytochrome b gene sequence of rice blast fungus was achieved by site‐directed mutagenesis. Molecular diagnostic methods were developed for identifying QoI resistance in rice blast fungus using the plasmid construct. CONCLUSION: As the management of rice blast disease is often dependent on chemicals, the rational design of control programmes requires a proper understanding of the fungicide resistance phenomenon in field populations of the pathogen. Mutation of the cytochrome b gene of rice blast fungus would be specifically detected from diseased leaves and seeds using the molecular methods developed in this study. Copyright © 2009 Society of Chemical Industry     

Effects of co‐supply ratios of swimming crab Portunus trituberculatus zoeae and Artemia on survival and growth of East Asian common octopus Octopus sinensis paralarvae under an upwelling culture system

Despite recent advances in culture technology for East Asian common octopus Octopus sinensis paralarvae using upwelling systems, securing suitable feed for the paralarvae is an unresolved issue. The zoea of the swimming crab Portunus trituberculatus is a good candidate for paralarval feed because of the high fecundity of the adult females. To investigate the effects of supplying P. trituberculatus zoeae and their feeding method on paralarvae, we cultured paralarvae with supplying different combination ratios of zoeae and Artemia (10:0, 7:3, 5:5, 3:7 and 0:10), and with or without supplementing rotifers using small‐scale (3‐L) upwelling systems. Paralarval survival rate and growth were improved when zoeae were supplied as the main feed, but reduced when the proportion of Artemia exceeded half the whole preys. Supplementing rotifers did not affect the paralarval survival and growth. Subsequently, paralarvae were cultured by supplying zoeae (partially augmented by Artemia) using three large (1‐kl) upwelling systems to assess their feeding effectiveness in juvenile octopus production. Paralarvae could be cultured at high survival rates of 77.1 ± 5.1% to reach benthic juveniles at 23 days after hatching. In conclusion, supplying P. trituberculatus zoeae augmented with Artemia under an upwelling culture system has great potential for juvenile octopus production.     

Effect of taurine supplementation on hepatic metabolism and alleviation of cadmium toxicity and bioaccumulation in a marine teleost,red sea bream, <Emphasis Type="Italic">Pagrus major</Emphasis>

This study was performed to unravel the mechanism of the beneficial action of taurine on marine teleost fish, red sea bream (Pagrus major), by analyzing the hepatic metabolism. Moreover, the ameliorative effects of the nutrient against cadmium toxicity and bioaccumulation were further evaluated. The fish were fed a diet containing 0 % (TAU0 %), 0.5 % (TAU0.5 %), or 5.0 % (TAU5.0 %) taurine for 40–55 days (d) and subjected to cadmium acute toxicity and bioaccumulation tests. Taurine deficiency in feed severely affected growth and the hepatic metabolic profiles of the fish, including a remarkable increase in myo-inositol, aspartate, and ß-alanine in the TAU0 % group, which indicates a complementary physiological response to taurine deficiency. For the acute toxicity test, fish were fed the test diets for 55 d and were then exposed to different dose of cadmium ranging from 0 to 5.6 mg/L for 96 h. Fish fed taurine had a higher tolerance to cadmium than those not fed taurine. For the bioaccumulation test, fish were fed the test diets for 40 d and then were chronically exposed to 0.2 mg/L of cadmium for 28 d followed by depuration for 21 d. Cadmium concentrations in the liver and muscle of fish fed TAU5.0 % were significantly lower than those of fish fed TAU0 % for the first 7 d of exposure and the first 7 d of elimination. Our findings suggest a possible mechanism for the beneficial role played by taurine and that the inclusion of taurine in fish aquaculture feed may reduce cadmium contamination of fish intended for human consumption.     

Alternaria brassicae produces a host-specific protein toxin from germinating spores on host leaves

Spore suspensions of Alternaria brassicae, the causal agent of gray leaf spot in Brassica plants, were incubated on the leaves of cabbage (B. oleracea) and spore germination fluid (SGF) was collected after 48 h. A high molecular weight (HMW) fraction (>10 kDa) was separated from the SGF by ultrafiltration. In a detached leaf assay, the HMW fraction induced visible symptoms only on host leaves and the toxicity was lost by treatment with proteinase K or heat at 60 degrees C for 15 min, indicating the presence of host-specific protein toxin(s). A protein toxin in the HMW fraction was purified by several chromatography steps. The toxin induced water-soaked symptoms followed by chlorosis at concentrations of 0.5 to 1 microg/ml on host leaves, but not on nonhost leaves even at 50 microg/ml. The toxin also had infection-inducing activity when added to spore suspension of a nonpathogenic isolate of A. alternata, causing symptoms similar to the infection of A. brassicae only on host leaves. These results indicate that a new host-specific protein toxin named ABR-toxin is released from germinating spores of A. brassicae on host leaves. ABR-toxin migrated as a protein of 27.5 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The isoelectric point of ABR-toxin was estimated to be approximately 7.0 and 21 N-terminal amino acid residues were sequenced.     

Toxicity of chlorothalonil to marine organisms

We evaluated the acute toxicity of the antifouling biocide 2,4,5,6-tetrachloro-1,3-benzenedicarbonitrile (chlorothalonil, TPN) to an alga (Skeletonema costatum), crustacea (Tigriopus japonicus, kuruma prawn Marsupenaeus japonicus), and a teleost fish (red sea bream Pagrus major) commonly found in the coastal waters of Japan. We also performed acute toxicity and early-life-stage tests for TPN using mummichog Fundulus heteroclitus and used the results to predict the chronic toxicity of TPN to red sea bream. The acute toxicity values of TPN (based on measured concentrations) for S.?costatum, T.?japonicus, M.?japonicus, red sea bream, and mummichog were 0.95, 16, 290, 35, and 61???g/l, respectively. The early-life-stage test with mummichog embryos revealed that hatchability and growth were the most sensitive indicators, and the lowest- and no-observed-effect concentrations, based on the measured concentrations, were 32 and 11???g/l, respectively. We estimated the chronic toxicity value for red sea bream to be 11???g/l based on these toxicity values. Considering the current measured concentrations of TPN in coastal waters of Japan, its environmental impact would appear to be limited.     

Early detection of urinary bladder carcinogens in rats by immunohistochemistry for γ-H2AX: a review from analyses of 100 chemicals

In safety evaluations of chemicals, there is an urgent need to develop short-term methods to replace long-term carcinogenicity tests. We have reported that immunohistochemistry for γ-H2AX, a well-established biomarker of DNA damage, can detect bladder carcinogens at an early stage using histopathological specimens from 28-day repeated-dose oral toxicity studies in rats. Given the markedly low level of γ-H2AX formation in the bladder urothelium of untreated rats, an increase in γ-H2AX-positive cells following chemical exposure can be relatively easy to identify. Among the 100 compounds examined to date, bladder carcinogens can be detected with high sensitivity (33/39; 84.6%) and specificity (58/61; 95.1%). As expected, γ-H2AX formation levels tended to be high following exposure to genotoxic bladder carcinogens, whereas nongenotoxic bladder carcinogens also increased the number of γ-H2AX-positive cells, probably through secondary DNA damage associated with sustained proliferative stimulation. γ-H2AX formation in the bladder urothelium reflects species differences in susceptibility to bladder carcinogenesis between rats and mice and shows a clear dose-dependency associated with the intensity of tumor development as well as high reproducibility. Some of the bladder carcinogens that showed false-negative results in the evaluation of γ-H2AX alone could be detected by combined evaluation with immunostaining for bladder stem cell markers, including aldehyde dehydrogenase 1A1. This method may be useful for the early detection of bladder carcinogens, as it can be performed by simple addition of conventional immunostaining using formalin-fixed paraffin-embedded tissues from 28-day repeated-dose toxicity studies in rodents, which are commonly used in safety evaluations of chemical substances.     

GnRH systems in masu salmon and barfin flounder

Multiple forms of the gonadotropin-releasing hormone (GnRH) exist in teleost fish. A salmonid fish, masu salmon Oncorhynchus masou has salmon GnRH (sGnRH) and chicken GnRH-II (cGnRH-II). sGnRH neurons were scattered from the olfactory nerve through the ventral telencephalon (VT) and the preoptic area (POA). sGnRH but not cGnRH-II was detected in the pituitary. sGnRH mRNA levels in the VT and the POA increased during gonadal maturation, suggesting that sGnRH neurons in these areas are involved in gonadal maturation. sGnRH neurons were first detected in a cluster near the olfactory epithelium 40 days after fertilization. sGnRH neurons were not detected in the brain by the olfactory epithelia lesion, suggesting that sGnRH neurons are derived from the olfactory epithelium. A pleuronectiform fish, barfin flounder Verasper moseri has sGnRH, cGnRH-II and seabream GnRH (sbGnRH). sGnRH and cGnRH-II-immunoreactive fibers were observed throughout the brain, but not in the pituitary. sbGnRH neurons were located in the POA and sent fibers to the pituitary, indicating that sbGnRH is involved in GTH secretion. Judging from the location of neuronal somata and their projections, it is indicated that three GnRH systems exist in the barfin flounder; the TN-, the MT- and the POA-GnRH system. However, in masu salmon, clear anatomical identification of the TN- and the POA-GnRH system is difficult, because the GnRH neurons located in the ventral forebrain are consecutive and the GnRH form produced in these neurons is the same (sGnRH). Thus, it is suggested in masu salmon that sGnRH neurons are derived from the olfactory epithelium, migrate into the brain and play different roles according to the location in the brain.