Authentication of flying-fish-meal content of processed food using PCR-RFLP

Nucleotide sequence and polymerase chain reaction (PCR)-restriction fragment length polymorphism (RFLP) analysis of the 3′-portion of the mitochondrial 16S RNA gene (rDNA) coding sequence were used to differentiate between flying fishes and other fishes that are frequently used in ago-noyaki production. In this study, we successfully distinguished between flying fishes and the other fishes by combining amplified DNA fragments with universally designed primers and digesting the PCR products with AfaI and MfeI restriction endonucleases. PCR-RFLP of 15 ago-noyaki, 2 noyaki, and 8 other processed flying-fish products were analyzed to authenticate flying-fish content among processed seafood products. After digestion of the PCR products with both enzymes, we found that all ago-noyaki and processed flying fish products contained either two or three DNA fragments of ~200, 300, and 530 bp, and noyaki samples (which do not contain flying fish) contained only one fragment of ~530 bp. Here, we present a new procedure to confirm the content of flying-fish meal in ago-noyaki.     

Plasma concentration and cardiovascular effects of lidocaine during continuous epidural administration in dogs anesthetized with isoflurane

The cardiovascular effects of continuous epidural administration (CEA) of lidocaine were investigated in anesthetized dogs. Loading epidural injections of 2, 4, or 6 mg/kg of lidocaine were followed by CEA with 1, 2, or 3 mg/kg/hr lidocaine, respectively, for 2 hr under 2.0% isoflurane anesthesia. Heart rate, direct blood pressure, cardiac index, and stroke volume decreased dose-dependently during CEA, whereas systemic vascular resistance did not significantly differ with dose, and no characteristic changes were observed in any groups. Plasma lidocaine concentration reached a steady state during CEA and increased in a dose-dependent manner. Circulatory suppression caused by lidocaine CEA was not attributable to peripheral vasodilation, but rather to the direct cardiac action of systemic lidocaine absorption from the peridural space.     

Finding of a highly efficient ZFN pair for Aqpep gene functioning in murine zygotes

The generation efficiencies of mutation-induced mice when using engineered zinc-finger nucleases (ZFNs) have been generally 10 to 20% of obtained pups in previous studies. The discovery of high-affinity DNA-binding modules can contribute to the generation of various kinds of novel artificial chromatin-targeting tools, such as zinc-finger acetyltransferases, zinc-finger histone kinases and so on, as well as improvement of reported zinc-finger recombinases and zinc-finger methyltransferases. Here, we report a novel ZFN pair that has a highly efficient mutation-induction ability in murine zygotes. The ZFN pair induced mutations in all obtained mice in the target locus, exon 17 of aminopeptidase Q gene, and almost all of the pups had biallelic mutations. This high efficiency was also shown in the plasmid DNA transfected in a cultured human cell line. The induced mutations were inherited normally in the next generation. The zinc-finger modules of this ZFN pair are expected to contribute to the development of novel ZF-attached chromatin-targeting tools.     

Comparison of the cardiovascular effects of intracellular cyclic adenosine 3',5'-monophosphate (cAMP)-modulating agents in isoflurane-anesthetized cats

The inotropic, chronotropic, and vasodilatory effects of five commonly used cyclic adenosine 3',5'-monophosphate (cAMP)-modulating agents were evaluated. Hemodynamic functions were measured continuously in isoflurane-anesthetized cats during infusion of the following: dobutamine (DOB; 2.5, 5 and 10 microg/kg/min; n=8), dopamine (DOP; 1.25, 2.5, 5 and 10 microg/kg/min; n=5), milrinone (MIL; 2.5, 5 and 10 microg/kg/min; n=8), 6-(3-dimethyl-aminopropionyl) forskolin hydrochloride (COL; 0.2, 0.4, 0.8, and 1.6 microg/kg/min; n=7), and bucladesine sodium (BUC; 10, 20, and 40 microg/kg/min; n=9). At the highest infusion rate, DOB and DOP produced the greatest positive inotropic (increase in left ventricular (LV) dP/dt = 89 +/- 4% and 75 +/- 6%, respectively) and chronotropic (increase in heart rate (HR) = 42 +/- 4% and 22 +/- 6%, respectively) effects. MIL and COL produced similar albeit less pronounced positive inotropic (increase in LV dP/dt = 18 +/- 3% and 22 +/- 6%, respectively) and chronotropic (increase in HR = 13 +/- 4% and 21 +/- 4%, respectively) effects. Both also had significant vasodilatory effects (decrease in peripheral resistance (PR) = -30 +/- 2% and -35 +/- 7%, respectively). In contrast, BUC produced only vasodilatation (decrease in PR = -33 +/- 6%). Hence, MIL, COL, and BUC had significant vasodilatory effects and less-pronounced inotropic effects than the catecholamines DOB and DOP. The vasodilatory effects of non-catecholamine drugs for treatment of congestive heart failure should translate into beneficial decreases in both pre-load and after-load. In contrast, the strong inotropic effects of DOB and DOP should be beneficial in the treatment of acute heart failure and anesthetic crisis.     

Estrogen modulates the skeletal muscle regeneration process and myotube morphogenesis: morphological analysis in mice with a low estrogen status

The purpose of this study was to elucidate the functions of estrogen and two estrogen receptors (ERs; ERα and ERβ) in the myoregeneration process and morphogenesis. Cardiotoxin (CTX) was injected into the tibialis anterior (TA) muscles of ovariectomized (OVX) mice to induce muscle injury, and subsequent myoregeneration was morphologically assessed. The diameter of regenerated myotubes in OVX mice was significantly smaller than that in intact mice at all time points of measurement. OVX mice also showed lower muscle recovery rates and slower speeds than did intact mice. ER protein levels showed a predominance of ERβ over ERα in both intact and OVX states. The ERβ level was increased significantly at 7 days after CTX injection in OVX mice and remained at a high level until 14 days. In addition, continuous administration of E2 to OVX mice in which muscle injury was induced resulted in a significantly larger diameter of regenerated myotubes than that in mice that did not receive estrogen. The results indicate that estrogen is an essential factor in the myoregeneration process since estrogen depletion delayed myoregeneration in injured muscles and administration of estrogen under the condition of a low estrogen status rescued delayed myoregeneration. The results strongly suggested that ERβ may be a factor that promotes myoregeneration more than does ERα.     

Risk of introduction of BSE into Japan by the historical importation of cattle from the United Kingdom and Germany

All cattle of UK and German origin imported to Japan since 1980 and slaughtered before 2002 were traced (n = 33 and 15 respectively) and the probability that none, one, two or three of these imported cattle had developed BSE (reached the end or last stage of incubation period) at the year of slaughter/death was calculated. The predicted risk that BSE was introduced into Japan by imported cattle was 0.18. Among cattle imported from these countries in various years, cattle imported from the UK in 1987 and 1988 presented the highest risk, while the risk that BSE entered Japan by live cattle imported from the UK in 1982 and from Germany in 1993 was negligible. Because there was no effective system to avoid the recycling of the BSE agent, those infected cattle imported from the UK in 1987 and 1988 most probably entered the feed chain in Japan in 1992 and 1993.     

Simulating the BSE epidemic and multiplication factor in dairy herds in Japan

With the objective of evaluating the effectiveness of an administrative guidance on the use of ruminant meat-and-bone meal in ruminant feed, effective from April 1996 to September 2001, we developed a model to simulate the evolution of the BSE epidemic and to estimate the BSE multiplication factor (K) in the Japanese dairy population. The output that provided the best fit to the number of BSE cases both observed and predicted to date suggest that the probability that bovine MBM was fed back to cattle was 14.2-75.2% and 0.129-0.570% during the periods from 1992 to April 1996 and from April 1996 to October 2001, respectively. Given these estimates, the value of K would have peaked in 1995 at 40-48 and then declined to 0.32-0.67 between 1997 and 2001. These results suggest that the administrative guidance was effective in reducing the amount of MBM fed to cattle by a factor of 104-141 and was perhaps enough to drive the epidemic towards extinction.     

Evaluation of antimicrobial use in dairy cattle,beef cattle and broilers in Japan using dosage-based indicators

The sales amount of antimicrobials intended for use in dairy cattle, beef cattle and broilers from 2008 to 2019 was evaluated for each antimicrobial class and administration route using dosage-based indicators. Our results revealed that the antimicrobial sales amount sold for use in dairy cattle in 2019 in terms of total weight of active ingredient, the number of defined daily doses (DDDs) (theoretical amount of biomass subjected to antimicrobial treatment in a year) and the number of treatment days (TDs) (theoretical number of days of treatment that an animal is subjected to in a year) calculated using Japanese DDD values (DDDjp values) was 36,751 kg, 8,261,848,000 kg·days and 15.5 days, respectively. Likewise, the antimicrobial sales amount sold for use in beef cattle and broilers in 2019 in terms of these metrics was 33,403 kg, 3,928,248,000 kg·days and 3.61 days, and 69,773 kg, 2,947,848,000 kg·days and 10.66 days, respectively. There was a considerable difference between the number of DDDs calculated using DDDjp values and that calculated using European DDD values (DDDvet values) for antimicrobial products sold for use in dairy and beef cattle. Our results also revealed that the sales amount of some antimicrobials, such as cephalosporins and quinolones represented larger proportions when calculated using dosage-based indicators than when calculated using the weight of active ingredient. Considering that Japanese veterinarians and farmers are more likely to conform to the Japanese dosage recommendations rather than the European ones, these results indicate the need for using dosage-based metrics, in particular metrics based on Japanese dosages rather than European dosages.     

Analysis of respiratory quinones in soil for characterization of microbiota

Method for the analysis of respiratory quinones in soil was developed to characterize soil microbiota. The respiratory quinones were extracted with a mixture of chloroform and methanol using a Wahling blender or a sonicator and cleaned-up by a silicagel column cartridge. The quinone species were determined by reverse-phase high performance liquid chromatography. Spectra of peaks were measured with a photodiode array detector to examine the purity. More than 90% of extractable quinones in soil were recovered by three extractions. The recovery of quinones, added to soil as freeze-dried powder of microorganisms, was higher than 96%. This procedure led to higher representative results as follows. Fluctuations within 95% of cumulative frequency were 13% for the extracted amount of quinones and 20% for the dissimilarity, respectively. The quinone profiles of four soils were determined as follows: a soil from the aerobic layer of a paddy field, an ando soil and two yellow upland soils which had received chemical fertilizers with and without farmyard manure. The quinone profiles displayed clear differences in the microbial composition and in the microbial diversity among the soils. It was demonstrated that the analysis of the respiratory quinone profile was useful to characterize the microbial community structure in soil.