Verification by polymerase chain reaction of vertical transmission of Theileria sergenti in cows

To evaluate the transplacental transfer of Theileria sergenti infection in cattle, we used DNA probes to detect T. sergenti in 6 pregnant cows and their calves. All the animals were monitored by parasitologic, serologic, and polymerase chain reaction (PCR) assays for a predicted 875-base-pair (bp) DNA product and a 684-bp amplicon detected by nested PCR in the blood and spleens of aborted fetuses. An open reading frame (ORF) starting at nucleotide 170 and terminating at position 1021 was shown to code for a polypeptide of 283 amino acid residues. All 6 dams and 5 calves were positive for T. sergenti in all tests. One calf was positive only with nested PCR. We conclude that transplacental transmission of T. sergenti is a significant problem. The relevance of the data in the programmed introduction of new (especially pregnant) animals into established clean herds needs serious consideration with regard to control of theileriosis and other tickborne diseases.     

Effects of Confluent, Roscovitine Treatment and Serum Starvation on the Cell-cycle Synchronization of Bovine Foetal Fibroblasts

The present study was designed to examine the effects of cell-cycle synchronization protocols, such as confluent, roscovitine treatment and serum starvation, in bovine foetal fibroblasts on synchronization accuracy at G0/G1, viability, apoptosis, necrosis and ploidy for use as a nuclei donor. The cells in 5-10 passages were randomly allocated into three treated groups. Cells were cultured either in Dulbecco's modified Eagle's medium (DMEM) + 10% foetal bovine serum (FBS) until 90% confluent (group 1, confluent), in DMEM + 10% FBS + 30 microM roscovitine for 12 h (group 2, roscovitine), or in DMEM + 0.5% FBS for 5 days (group 3, serum starvation). Most of the cells (>80%) in all groups were arrested at the G0/G1 stage. Although the rates did not differ, cells in group 1 showed an increased cell population arrested at the G0/G1 phase. Significantly (p < 0.05) higher rates of apoptosis occurred in group 3 than in group 1 and 2 (10% vs 6% and 6%, respectively). No differences in chromosomal abnormality were observed among groups. However, by increasing the number of cell culture passages up to 15, significantly (p < 0.05) higher chromosomal abnormality was observed than in 5 and 10 passages (39% vs 28% and 23%, respectively) in group 1. The results clearly indicated that bovine foetal fibroblasts could be effectively synchronized at G0/G1 stages by all the three different treatments, confluent, roscovitine and serum starvation. However, cells in confluent showed reduced apoptosis and necrosis when they underwent 5-10 passages, exhibiting increased percentage of cells with stable chromosome diversity. Hence, cells in confluent merit further studies before they could be used as nuclear donors.     

Antihypertensive Effects of IGTGIPGIW Peptide Purified from Hippocampus abdominalis: p-eNOS and p-AKT Stimulation in EA.hy926 Cells and Lowering of Blood Pressure in SHR Model

The aim of this study was to assess the potential hypertensive effects of the IGTGIPGIW peptide purified from Hippocampus abdominalis alcalase hydrolysate (HA) for application in the functional food industry. We investigated the antihypertensive effects of IGTGIPGIW in vitro by assessing nitric oxide production in EA.hy926 endothelial cells, which is a major factor affecting vasorelaxation. The potential vasorelaxation effect was evaluated using 4-amino-5-methylamino-2′,7′-difluorofluorescein diacetate, a fluorescent stain. IGTGIPGIW significantly increased the expression of endothelial-derived relaxing factors, including endothelial nitric oxide synthase and protein kinase B, in EA.hy926 cells. Furthermore, oral administration of IGTGIPGIW significantly lowered the systolic blood pressure (183.60 ± 1.34 mmHg) and rapidly recovered the diastolic blood pressure (143.50 ± 5.55 mmHg) in the spontaneously hypertensive rat model in vivo. Our results demonstrate the antihypertensive activity of the IGTGIPGIW peptide purified from H. abdominalis and indicate its suitability for application in the functional food industry.     

DNA profiling and genetic diversity of Korean soybean (<Emphasis Type="Italic">Glycine max</Emphasis> (L.) Merrill) landraces by SSR markers

Approximately 7,000 accessions of Korean soybean (Glycine max (L.) Merrill) landraces, largely composed of three collections, the Korea Atomic Energy Research Institute’s soybean (KAS), the Korean Crop Experiment Station’s soybean (KLS) and the Korean Agricultural Development and Technology Center’s soybean (KADTC) collections, have been conserved at the Rural Development Administration (RDA) genebank in Korea. The accessions within collections were classified based on their traditional uses such as sauce soybean (SA), sprouted soybean (SP), soybean for cooking with rice (SCR), and OTHERS. A total of 2,758 accessions of Korean soybean landraces were used to profile and to evaluate genetic structure using six SSR loci. A total of 110 alleles were revealed by at the six SSR loci. The number of alleles per SSR locus ranged from 9 to 39 in Satt187 and Satt_074, respectively. The number of alleles ranged from 87 in the KADTC collection to 96 in the KLS collection, and from 63 in the SCR group to 95 in the SP group. Nei’s average genetic diversity ranged from 0.68 to 0.70 across three collections, and 0.64 to 0.69 across the usage groups. The average between-group differentiation (G _st) was 0.9 among collections, and 4.1 among the usage groups. The similar average diversity among three collections implies that the genetic background of the three collections was quite similar or that there were a large number of duplicate accessions in three collections. The selection from the four groups classified based upon usage may be a useful way to select accessions for developing a Korean soybean landrace core collection at the RDA genebank. DNA profile information of accessions will provide indications of redundancies or omissions and aid in managing the soybean collection held at the RDA genebank. The information on diversity analysis could help to enlarge the genetic diversity of materials in breeding programs and could be used to develop a core collection.     

Migration of Strongyloides venezuelensis in rats after oral inoculation of free-living infective larvae

Strongyloides venezuelensis (SVZ) infection was chronologically monitored in 85 Sprague-Dawley rats (SDR), which were orally inoculated with approximately 1,000 infective larvae. In order to describe the characteristics of migrating larvae (MLS) in various visceral organs (the liver, lung, cardiac blood, and small intestine), 5 SDR were sacrificed at 20 min, 45 min, 1 hr, 2 hr, 3 hr, 4 hr, 8 hr, 12 hr, 16 hr, 48 hr, 72 hr, 96 hr, 120 hr, 144 hr, 168 hr and 192 hr post inoculation (PI). MLS were recovered from the liver and blood 20 and 45 min PI and measured 788 +/- 26 microm and 846 +/- 40 microm in length, respectively. MLS were first observed in the lung tissue 45 min PI and measured 925 +/- 38 microm on the average. In the trachea, MLS measuring 849 +/- 75 microm appeared 3 to 96 hrs PI. Adult worms (AWS) measuring 1,926 +/- 521 microm to 2,956 +/- 159 microm in length were observed in the small intestine from 120 hr PI. The worms appeared to mature more than 168 hr PI and attained the average maximum length of 2,420 +/- 532 microm. At 3 hr PI focal hyperemic and necrotic lesions were evidently observed in the liver and lung, together with eosinophilic infiltration in the stomach, liver, and lung. The parasites were histologically detectable in the lung tissues but were very difficult to find in the liver and the epithelial layer of small intestine. These data demonstrate that SVZ parasites take 20 min to reach the liver via the stomach and only three hours to reach the trachea through the same route. The development from eggs to adults takes 168 hr in the SDR model.     

Protective effect of butylated hydroxylanisole against hydrogen peroxide-induced apoptosis in primary cultured mouse hepatocytes

Butylated hydroxyanisole (BHA) is a synthetic phenolic compound consisting of a mixture of two isomeric organic compounds: 2-tert-butyl-4-hydroxyanisole and 3-tert-butyl-4-hydroxyanisole. We examined the effect of BHA against hydrogen peroxide (H₂O₂)-induced apoptosis in primary cultured mouse hepatocytes. Cell viability was significantly decreased by H₂O₂ in a dose-dependent manner. Additionally, H₂O₂ treatment increased Bax, decreased Bcl-2, and promoted PARP-1 cleavage in a dose-dependent manner. Pretreatment with BHA before exposure to H₂O₂ significantly attenuated the H₂O₂-induced decrease of cell viability. H₂O₂ exposure resulted in an increase of intracellular reactive oxygen species (ROS) generation that was significantly inhibited by pretreatment with BHA or N-acetyl-cysteine (NAC, an ROS scavenger). H₂O₂-induced decrease of cell viability was also attenuated by pretreatment with BHA and NAC. Furthermore, H₂O₂-induced increase of Bax, decrease of Bcl-2, and PARP-1 cleavage was also inhibited by BHA. Taken together, results of this investigation demonstrated that BHA protects primary cultured mouse hepatocytes against H₂O₂-induced apoptosis by inhibiting ROS generation.     

Temperature-dependent development and survival of Podisus maculiventris (Hemiptera: Pentatomidae): implications for mass rearing and biological control

The spined soldier bug, Podisus maculiventris (Say), is a generalist predator that can control >75 insect pests in various agroecosystems. Despite valuable results from the previous studies, thermal development and survival of P. maculiventris over a wide range of temperatures had not been elucidated completely. This study was conducted to investigate the stage-specific development and survival of P. maculiventris at eight constant temperatures (13.2, 18.4, 21.7, 23.7, 27.2, 32.7, 35.2, and 40.6 °C) covering the entire thermal range for P. maculiventris development. Completion of egg development was observed at 13.2–32.7 °C, whereas nymphs successfully developed into adults at 18.4–32.7 °C. Survival model using log-normal equations showed bell-shape patterns for all stages, and temperatures resulting in highest survival of P. maculiventris were 19.9, 24.3, and 24.5 °C for egg, nymph, and egg to adult, respectively. Developmental rates at the eight temperatures were fitted with a nonlinear Briere model, which estimated optimal temperatures for development as 31.2, 30.6, and 30.6 °C for egg, nymph, and egg to adult, respectively. Operative thermal ranges (i.e., in-between the lower and upper developmental thresholds) were estimated to be 8.9–35.2, 12.8–35.2, and 12.7–35.2 °C for egg, nymph, and egg to adult, respectively. In a linear model, the lower thresholds were 10.9, 13.1, and 13.0 °C for egg, nymph, and egg to adult, respectively. Findings herein provide comprehensive data and explanations on optimal temperature and thermal requirement for P. maculiventris, which can be used to optimize environmental condition in mass rearing and predicting phenology of P. maculiventris in the field.     

Effect of in vitro xenoestrogens on steroidogenesis in mature female fish, Chasmichthys dolichognathus

In this study, fully vitellogenic oocytes of the longchin goby (Chasmichthys dolichognathus) were exposed to in vitro xenoestrogens such as diethylstilbestrol (DES), bisphenol A (BPA) and nonylphenol (NP) at concentrations of 100 ng/ml in the presence of [³H]17α-hydroxyprogesterone as precursor. The major metabolites produced in vitro are progestogens [17α-hydroxy,20α-dihydroprogesterone (17α20αOHP) and 17α-hydroxy,20β-dihydroprogesterone (17α20βOHP)], androgens [androstenedione (A4) and testosterone (T)] and estrogens [estrone (E1) and estradiol-17β (E2)]. Comparative activities of these chemicals in oocyte steroid biosynthesis showed as follows: NP treatment resulted in clear stimulation of estrogen production, while the opposite occurred in DES treated incubation. Treatment with DES resulted in a higher synthesis of 17α20αOHP. BPA treatment increased the androgen, while estrogen synthesis was slightly inhibited. These results suggest that NP exhibited clearly estrogenic activity on the three chemicals.