Accumulation of paralytic shellfish poisoning toxin by the swimming crab <Emphasis Type="Italic">Charybdis japonica</Emphasis> in Kure Bay,Hiroshima Prefecture

The accumulation of paralytic shellfish poisoning (PSP) toxin in the crab Charybdis japonica was investigated in Kure Bay, when a bivalve Mytilus galloprovincialis, in the same water, showed toxicity caused by PSP toxin. In 2005, 17 specimens of the crab had PSP toxin over 4 MU/g in the hepatopancreas, and the highest toxicity was at 37.4 MU/g. Since the regulation limit of PSP for crab hepatopancreas was set in 2004, this is the first observation of regulatory level of PSP toxin in C. japonica. Unlike in the hepatopancreas of the crab, the hazardous level of the toxin was not detected in the muscle of the cephalothorax and the appendage. The toxin accumulation in the crab was also investigated by feeding toxic mussels to the crab. The crab retained the toxin mainly in the hepatopancreas, and the ratio of retention in the crab was from 12.9 to 24.6%. The toxin profiles, shown in the feeding experiments, suggest that the transformation of PSP toxin occurs in the crab because dcGTX2 and dcGTX3 was detected in all crab specimens despite the lack of these analogs in the mussels used as feed.     

A step forward in development of fish protein hydrolysate-based diets for larvae of Japanese eel Anguilla japonica

Eggs of spiny dogfish Squalus acanthias are an indispensable and effective component of the compound diets currently used for larvae of the Japanese eel Anguilla japonica. However, this fish species, a natural resource, is becoming more scarce, suggesting that suitable diets should be developed using more sustainable components. In the search for appropriate diets without shark eggs, we have designed test diets in which shark eggs were replaced with general feed materials, such as fish protein hydrolysate (FPH), with the notable improvement that the FPH had been pre-digested with integral enzymes from frozen krill. Feeding trials were conducted to assess the effects of the test diets on survival and growth of Japanese eel larvae at early developmental stages. Larvae fed on the test diets were found to be able to survive up to 65 days after hatching, the maximum total length and body depth were 14.1 and 1.5 mm, respectively. Although larvae fed on the test diets still exhibited a low survival rate and poor growth, the FPH-based diets had a nutritional value that promoted substantial larval growth based on the significant difference between the initial and final body sizes. Our results suggest that the development of FPH-based diets not based on shark eggs but rather on general feed ingredients is promising but that there is much room for improvement.     

Quantification of lipophilic toxins associated with diarrhetic shellfish poisoning in Japanese bivalves by liquid chromatography-mass spectrometry and comparison with mouse bioassay

ABSTRACT:   Quantification of lipophilic toxins in bivalves associated with diarrhetic shellfish poisoning was investigated by liquid chromatography–mass spectrometry (LC–MS). Using a C8-silica reversed phase column and a mobile phase of aqueous acetonitrile containing 2 mM ammonium formate and 50 mM formic acid, okadaic acid, dinophysistoxin-1, 7- O -palmitoyldinophysistoxin-1, pectenotoxin-1, pectenotoxin-2, pectenotoxin-6, pectenotoxin-2 seco-acid, yessotoxin, and 45-hydroxyyessotoxin in bivalves were quantified by LC–MS in the negative mode. When the crude 90% methanol extracts were analyzed by LC–MS, there were no significant effects from bivalve matrices on the quantification of toxins. More than 200 bivalve samples collected from various production areas in Japan were analyzed by LC–MS. Pectenotoxin-6 and dinophysistoxin-1 were the dominant toxins in scallops and mussels, respectively. Yessotoxin and 45-hydroxyyessotoxin were also detected in both species. Comparison of the quantitative results obtained for these bivalve samples between LC–MS and mouse bioassay indicates that LC–MS is suitable for routine monitoring of lipophilic toxins in Japanese bivalves.     

Spawning induction and seed production of Eastern little tuna,Euthynnus affinis (Cantor, 1849), in the post‐ and pre‐spawning seasons by hormonal treatment in a semi‐closed recirculation system with elevated temperature

We previously established a method for spawning induction in Eastern little tuna (ELT) Euthynnus affinis (Cantor, 1849) by administering a gonadotropin‐releasing hormone analog (GnRHa) during the natural spawning season in Japan (August–October). In order to establish seed production of ELT in the off‐spawning season, we first conducted three spawning induction trials by GnRHa administration from October 2011 to January 2012 using ELT broodstock (2 years old; three females and four males) maintained in a 10‐m³ tank with a semi‐closed recirculation system and static elevated temperature. Average water temperature and daily egg production in three trials lasting 11–15 days were 27.0 ± 0.09°C and 268 173 eggs (Trial 1), 27.0 ± 0.11°C and 277 9098 eggs (Trial 2), and 25.5 ± 0.39°C and 291 113 eggs (Trial 3) respectively. Mean fertilization rate and mean hatching rate were 70.4% and 60.5% (Trial 1), 83.9% and 79.6% (Trial 2), and 62.5% and 57.4% (Trial 3) respectively. We also succeeded in producing ELT larvae in the pre‐spawning season (April–July), although the quantity and quality of larvae produced were inferior to those produced in other calendar months. In trials involving periodic GnRHa administration during the off‐spawning seasons, hatched larvae were obtained in the 10‐m³ tank after six of nine administrations in the 2011–2012 off‐spawning season and in 16 of 19 administrations in the 2012–2013 off‐spawning season. The findings of this study demonstrated that hormonal treatment and thermal control could be used to extend the spawning period in ELT, potentially allowing larval production in the post‐ and pre‐spawning seasons.     

A simulation study for a linear measurement error model when error variances vary between measurements

When predicting scores in the Draize eye irritation test based on measurements of in vitro alternative tests, we are often faced with estimating parameters in a linear measurement error model with heterogeneous error variances. This article proposes a new statistical method for parameter estimation to address this issue. The proposed method is an extension of an earlier proposal that applied a linear measurement error model with homogeneous error variances, to cases with heterogeneous error variances. A simulation study to examine the performance of the proposed method was conducted in a framework that was adaptable to the data, which was obtained in a validation study of alternative methods to animal experiments conducted in Japan. The proposed method reduced the biases of estimates in comparison with an ordinary regression analysis method and three other methods under the assumption of homogeneous error variances. Although the proposed method did not fit the real data well, the resulting prediction formula was far better than those obtained by other methods.     

Clearance of Theileria sergenti-infected bovine red blood cells in severe combined immune deficiency mice

Clearance of Theileria sergenti-infected bovine red blood cells (Bo-RBCs) from the blood circulation of severe combined immune deficiency (SCID) mice was studied to help understand the mechanisms of anemia developing in cattle infected with T. sergenti. For the clearance test, Bo-RBC samples having 2%, 58%, and 76% parasitemia and, as a control, parasite-free Bo-RBCs were prepared in the Bo-RBC-SCID mouse model. The T. sergenti-infected Bo-RBCs and the uninfected control Bo-RBCs were separately labeled with two, green and red, fluorescent dyes, mixed together, and injected intravenously into SCID mice. The blood samples collected at various time points were observed under a fluorescent microscope, and the numbers of green and red fluorescing RBCs were counted differentially to determine the clearance rates of T. sergenti-infected and uninfected Bo-RBCs. This test clearly demonstrated that the Bo-RBC samples having higher parasitemias were cleared faster from the blood circulation of SCID mice. The results suggest that the intravascular clearance system in SCID mice may have a mechanism by which T. sergenti-parasitized and non-parasitized Bo-RBCs are recognized and cleared differentially.     

低分子量热激蛋白Bmhsp19.9基因的表达

利用RT-PCR技术，对家蚕(Bombyx moil)低分子量热激蛋白Bmhsp19．9基因进行了定量表达分析。Bmhsp19．9在各组织中均有一定量的表达，但不同组织间的表达差异达10多倍，表达量丰富的组织是生长发育旺盛的组织，如精巢和卵巢，然后是丝腺、蛹等组织。其应激表达模式可分为3类，以其在精巢、后部丝腺和脂肪体中的强应激表达模式为主，经热刺激处理后其表达量有显著增加。经刺激诱导2h后其表达量达到最高，随后逐渐降低，在热刺激诱导12h后是未刺激的2～3倍，而此时Bmhsp19．9在脂肪体中的应激表达是其对照的10多倍。根据Bmhsp19．9在不同组织中的基本表达量、应激表达量和应激表达的持续时间各不相同，显示其与细胞分裂与分化等生命活动有一定联系。并认为其在后部丝腺、脂肪体、精巢、卵巢与血液中作用方式不一样，推测其作为分子伴侣在组织间的作用对象与作用效应也应有所差异。     

Effect of a commercial microbiological additive on beef manure compost in the composting process

We observed the effects of a commercial microbiological additive (MA) on the beef manure composting process and investigated the changes in its physical and chemical components and microbial populations in the composting process. When beef manure compost was inoculated with the MA, its temperature rapidly increased at the beginning of the process, and after the first turning, the ammonia emission from the compost pile and nitrate production decreased quicker than that in the composting process without the MA. During the composting process, the manure inoculated with MA showed a larger numbers of mesophilic and thermophilic aerobes, and a smaller number of thermophilic anaerobes than in the process without the MA. These results suggested that mesophilic aerobes increased in number and metabolized ammonia by assimilation rather than nitrification to accelerate the temperature elevation in the composting process with the MA.