Effect of intramammary injection of rboGM-CSF on milk levels of chemiluminescence activity, somatic cell count, and Staphylococcus aureus count in Holstein cows with S. aureus subclinical mastitis

The effect of intramammary injection of recombinant bovine granulocyte-macrophage colony-stimulating factor (rboGM-CSF, 400 microg/10 mL) on quarter milk levels of chemiluminescence (CL) activity, and somatic cell count (SCC) and shedding pattern of Staphylococcus aureus was investigated. Ten Holstein cows, naturally infected with S. aureus were used, with either early-stage or late-stage subclinical mastitis. Injection of rboGM-CSF caused a remarkable increase in milk CL activity with a peak at 6 h after the cytokine injection in the early- and late-stage groups. In the early-stage group, milk SCC stayed around preinjection level at 6 h, rose significantly on days 1 and 2, and was followed by a smooth and significant decline to an under preinjection level (below 200 000 cells/mL) on day 7 postinjection. Alternatively, in the late-stage group, milk SCC rose significantly at 6 h after the cytokine injection and maintained high levels thereafter. The milk S. aureus count decreased drastically by the cytokine injection in the early-stage group. The bacterial count was moderately decreased in the late-stage group, but increased back to preinoculation levels on day 7 after the cytokine injection. The results suggest that the rboGM-CSF has a potential as a therapeutic agent for S. aureus infection causing subclinical mastitis of dairy cows, if the cytokine is applied at the initial stage of infection.     

Sward characteristics,nutritive value and choice by cattle of conterminous monocultures of centipedegrass (Eremochloa ophiuroides) and bahiagrass (Paspalum notatum)

In south‐western Japan, centipedegrass (Eremochloa ophiuroides; CG) offers a novel option for a warm‐season perennial for grazing use in areas where bahiagrass (Paspalum notatum; BG) can be grown. However, the potential of CG as a forage has not been fully explored because of the short history as a forage crop. We conducted four experiments to evaluate CG (cv. TifBlair) in comparison with BG (cv. Pensacola) in terms of sward characteristics, nutritive value and choice by animals. In each experiment, four Japanese Black cows (Bos taurus) were individually allowed to graze conterminous monocultures of CG and BG (5 × 10 m each) for 30 min. Irrespective of regrowth durations and fertilizer rates, CG was consistently shorter, leafier and denser, contained lower acid detergent fiber and cellulose, and was preferred or equally selected by cows, as compared with BG. Furthermore, CG maintained sufficient levels of crude protein (80–89 g/kg DM) to ensure voluntary intake of ruminant animals under extended regrowth^? and without fertilizer, whereas BG failed to do so (65 g/kg DM). CG provided higher digestible dry matter than BG when crude protein concentration exceeded 86 g/kg DM. The results indicate advantages of CG as a forage.     

Comparison of dendritic cell-mediated immune responses among canine malignant cells

Dendritic cell (DC) vaccination is one of the most attractive immunotherapies for malignancies in dogs. To examine the differences in DC-mediated immune responses from different types of malignancies in dogs, we vaccinated dogs using autologous DCs pulsed with keyhole limpet hemocyanin (KLH) and cell lysate prepared from squamous cell carcinoma SCC2/88 (SCC-KLH-DC), histiocytic sarcoma CHS-5 (CHS-KLH-DC), or B cell leukemia GL-1 (GL-KLH-DC) in vitro. In vivo inductions of immune responses against these tumor cells were compared by the delayed-type hypersensitivity (DTH) skin test. The DTH response against SCC2/88 cells were observed in dogs vaccinated with autologous SCC-KLH-DC, while the response was undetectable against CHS-5 and GL-1 cells in dogs vaccinated with autologous CHS-KLH-DC and GL-KLH-DC. Skin biopsies taken from DTH challenge sites were then examined for immunohistochemistry, and recruitment of CD8 and CD4 T cells was detected at the site where SCC2/88 cells were inoculated in dogs vaccinated with SCC-KLH-DC. By contrast, neither CD8 nor CD4 T cell infiltration was found at the DTH challenge site in the dogs vaccinated with CHS-KLH-DC or GL-KLH-DC. These findings may reflect that the efficacy of immune induction by DC vaccination varies among tumor types and that immune responses could be inducible in squamous cell carcinoma. Our results encouraged further investigation of therapeutic vaccination for dogs with advanced squamous cell carcinoma in clinical trials.     

Synthesis and Structure–Activity Relationships of Miticidal 4,5-Dihydropyrazole-5-thiones

A series of novel 4,5-dihydropyrazole-5-thiones (DHPs) was synthesised by treating the corresponding dihydropyrazolones with ‘Lawesson’s reagent and evaluated for miticidal activity against two-spotted spider mites (Tetranychus urticae Koch). Of these, 3-(4-chlorophenyl)-4,4-dimethyl-1-phenyl-4,5-dihydropyrazole-5-thione, 3-(4-chlorophenyl)-4-ethyl-4-methyl-1-phenyl-4,5-dihydropyrazole-5-thione, 3-(4-chlorophenyl)-1-phenyl-4,5-dihydropyrazole-5-thione-4-spirocyclopentane and 4,4-dimethyl-1-phenyl-3-(4-trifluoromethyl-phenyl)-4,5-dihydropyrazole-5-thione were highly active (pEC₅₀>4·0) and were more effective than the miticide dicofol (pEC₅₀=3·879), which has traditionally been used for the control of phytophagous mites. Structure–activity relationship (SAR) studies were performed on each position of the pyrazole ring of DHPs. The results indicated that the unsubstituted phenyl, 4-substituted phenyl and thioxo groups on the 1-, 3- and 5-positions of DHPs respectively were required for activity. Quantitative SAR studies using physicochemical parameters of substituents and the capacity factor k′ as a hydrophobicity index suggested that: (a) the activities of all types of DHPs examined were mainly dominated by hydrophobicity, (b) the bulkiness of 4-substituents of the 3-phenyl ring favoured the activity and (c) the log k′ optimum for all DHPs was 1·675, equivalent to a log P_ow value of c. 5·0.     

Effects of Yuzu (Citrus junos) Peel from Waste as an Aquaculture Feed Supplement on Growth,Environmental Load,and Dark Muscle Discoloration in Yellowtail Seriola quinqueradiata

Beneficial utilization of yuzu (Citrus junos) peel was investigated to improve the commercial value of cultured yellowtail fish. The antioxidant effects of grated yuzu peel from pomace (yuzu paste) on prevention of dark muscle discoloration during storage in sliced yellowtail fish were tested. Four diets were prepared, containing 0, 10, 50, and 100 g yuzu paste in a 1,450-g diet (wet basis). Fish were fed one of the four diets for 10 weeks. Growth performance was not significantly different among the dietary groups, but fish fed a diet containing 100-g yuzu paste showed the least average body weight and a significant increase in fecal amount and fat levels in the feces, which suggest low digestibility and low absorption. Dark muscle discoloration was significantly reduced in all yuzu dietary groups compared to the control group during storage at 4°C. The optimal supplementation of yuzu paste in the fish diet is very effective at maintaining fish flesh freshness as a finishing diet without causing additional phosphorus or nitrogen pollution of the sea and inhibiting fish growth. This study provides a new way to utilize waste yuzu peel, which also helps reduce waste from yuzu processing.     

Artificially induced tetraploid masu salmon have the ability to form primordial germ cells

Diploid gametes generated with tetraploid animals are a stepping stone to improving chromosome manipulation techniques. However, artificially induced tetraploid individuals generally die soon after hatching. Diploid gametes could be induced by in vivo cultures of tetraploid primordial germ cells (PGCs) through germ-line chimera. In the present study, characteristics of PGCs were studied in inviable tetraploid masu salmon, Oncorhynchus masou. Histological observation of tetraploid embryos revealed that the same or smaller numbers of PGCs were observed and they migrate into the genital ridges as did diploid PGCs during gonadogenesis. By whole-mount in situ hybridization using vasa messenger RNA (mRNA), 4–35 vasa-positive signals were detected in a pair of genital ridges of tetraploids. By cytological observation of genital ridge cell suspensions, several large round cells were observed, some of which extended pseudopodia. They also contained large nuclei and round granules in their cytoplasm, characteristics of PGCs. As the results suggest that inviable artificial tetraploids have PGCs, we expect to achieve diploid gamete production through surrogate propagation and tetraploid fish production.     

Effects of dietary phosphorus level on non-faecal phosphorus excretion from yellowtail (Seriola quinqueradiata Temminck & Schlegel) fed purified and practical diets

Non-faecal phosphorus (P) was determined for large yellowtail to estimate a minimum available P requirement (Experiment  1) and to justify inorganic P supplementation in a fish meal-based diet (Experiment 2). In Experiment 1, purified diets with incremental P concentrations were fed to yellowtail (mean weight 917 g) at a feeding rate of 1.5% of body weight. The peaks of non-faecal P excretion appeared 5–6 h after feeding in fish fed more than 4.5 g available P kg⁻¹ dry diet. Broken-line analysis indicated that the minimum available P requirement was 4.4 g kg⁻¹ dry diet. In Experiment 2, a purified diet (PR) containing 6.5 g available P kg⁻¹ and a fish meal-based diet with (F1) and without (F0) additional phosphorus were fed to yellowtail (mean weight 1.1 kg) at 1.5% (PR) and 2% (F0 and F1) feeding rates respectively. There was no significant difference in P excretion between fish fed the F0 (5.5 g soluble P kg⁻¹ dry diet) and the PR diet. However, significantly higher (34.5%) amounts of non-faecal P excretions (7.4 g soluble P kg⁻¹ dry diet) were found in fish fed F1 compared with the F0 diet. This suggested that there was an excess of dietary P in the F1 diet and that supplementation is not needed in fish meal-based diets for large yellowtail.     

Study of the Structure–Activity Relationship of an Anti-Dormant Mycobacterial Substance 3-(Phenethylamino)Demethyl(oxy)aaptamine to Create a Probe Molecule for Detecting Its Target Protein

The current tuberculosis treatment regimen is long and complex, and its failure leads to relapse and emergence of drug resistance. One of the major reasons underlying the extended chemotherapeutic regimen is the ability of Mycobacterium tuberculosis to attain a dormant state. Therefore, the identification of new lead compounds with chemical structures different from those of conventional anti-tuberculosis drugs is essential. The compound 3-(phenethylamino)demethyl(oxy)aaptamine (PDOA, 1), isolated from marine sponge of Aaptos sp., is known as an anti-dormant mycobacterial substance, and has been reported to be effective against the drug resistant strains of M. tuberculosis. However, its target protein still remains unclear. This study aims to clarify the structure–activity relationship of 1 using 15 synthetic analogues, in order to prepare a probe molecule for detecting the target protein of 1. We succeeded in creating the compound 15 with a photoaffinity group that retained antimicrobial activity, which proved to be a suitable probe molecule for identifying the target protein of 1.     

我国三种野生泥鳅遗传多样性的微卫星分析

本研究采用5个微卫星分子标记技术对我国黑龙江泥鳅(M.mohoity)、北方泥鳅(M.bipartitus)、泥鳅(M.anguillicaudatus)进行了遗传多样性分析。结果表明,三种泥鳅的5个微卫星基因座共检测到等位基因数27个,微卫星基因座的等位基因数范围为3~7个。5个微卫星基因座在三种泥鳅的多态信息含量都达到了高度多态的水平,平均PIC值在0.526~0.722之间,说明三种泥鳅具有丰富的遗传多样性。黑龙江泥鳅、北方泥鳅和泥鳅的观测杂合度Ho分别为0.978,0.889和0.742,期望杂合度He分别为0.683、0.781、0.721。三种泥鳅的遗传距离为0.189~0.559,泥鳅和北方泥鳅群体间遗传距离最小为0.189,亲缘关系较近,而泥鳅和黑龙江泥鳅群体间遗传距离最大为0.559,亲缘关系较远。Hardy-Weinberg平衡检验P值。得出所有个体在5个微卫星位点上都无偏离现象。聚类结果显示黑龙江泥鳅、北方泥鳅、泥鳅群体分为二支,泥鳅和北方泥鳅聚为一类,之后两者再与黑龙江泥鳅聚为一类。