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.     

Effects of Dietary Supplementation of Barodon on Growth Performance,Innate Immunity and Disease Resistance of Juvenile Olive Flounder,Paralichthys olivaceus,Against Streptococcus iniae

This study was conducted to evaluate the effects of dietary supplementation of Barodon, an anionic alkali mineral complex, on growth, feed utilization, humoral innate immunity and disease resistance of olive flounder. A basal experimental diet was used as a control and supplemented with 0.1, 0.2, 0.3, 0.4, or 0.5% Barodon. Triplicate groups of fish (26.4 ± 0.2 g) were fed one of the diets to apparent satiation twice daily for 10 wk. The growth performance was enhanced (P < 0.05) linearly and quadratically in fish fed diets containing Barodon compared with that in fish fed the control. Feed utilization was significantly improved by Barodon supplementation. Serum lysozyme and antiprotease activities were increased quadratically in Barodon fed groups. Also, significantly higher superoxide dismutase activity was found in Barodon‐fed fish. Dietary supplementation of 0.1–0.3% Barodon resulted in significant enhancement of fish disease resistance against Streptococcus iniae. The findings in this study indicate that dietary supplementation of Barodon can enhance growth, feed utilization, innate immunity, and disease resistance of olive flounder and that the optimum level seems to be 0.1% in diets.     

Rapidly quantitative detection of Nosema ceranae in honeybees using ultra-rapid real-time quantitative PCR

BackgroundThe microsporidian parasite Nosema ceranae is a global problem in honeybee populations and is known to cause winter mortality. A sensitive and rapid tool for stable quantitative detection is necessary to establish further research related to the diagnosis, prevention, and treatment of this pathogen.ObjectivesThe present study aimed to develop a quantitative method that incorporates ultra-rapid real-time quantitative polymerase chain reaction (UR-qPCR) for the rapid enumeration of N. ceranae in infected bees.MethodsA procedure for UR-qPCR detection of N. ceranae was developed, and the advantages of molecular detection were evaluated in comparison with microscopic enumeration.ResultsUR-qPCR was more sensitive than microscopic enumeration for detecting two copies of N. ceranae DNA and 24 spores per bee. Meanwhile, the limit of detection by microscopy was 2.40 × 10⁴ spores/bee, and the stable detection level was ≥ 2.40 × 10⁵ spores/bee. The results of N. ceranae calculations from the infected honeybees and purified spores by UR-qPCR showed that the DNA copy number was approximately 8-fold higher than the spore count. Additionally, honeybees infected with N. ceranae with 2.74 × 10⁴ copies of N. ceranae DNA were incapable of detection by microscopy. The results of quantitative analysis using UR-qPCR were accomplished within 20 min.ConclusionsUR-qPCR is expected to be the most rapid molecular method for Nosema detection and has been developed for diagnosing nosemosis at low levels of infection.     

Effect of Dark,Hard, and Vitreous Kernel Content on Protein Molecular Weight Distribution and on Milling and Breadmaking Quality Characteristics for Hard Spring Wheat Samples from Diverse Growing Regions

Kernel vitreousness is an important grading characteristic for segregation of subclasses of hard red spring (HRS) wheat in the United States. This research investigated the protein molecular weight distribution (MWD) and the flour and baking quality characteristics of different HRS wheat market subclasses. The U.S. regional crop quality survey samples obtained from six regions for three consecutive growing years were used for subclass segregation based on the dark, hard, and vitreous (DHV) kernel percentage. Flour milled from HRS wheat with greater percentages of DHV kernel showed higher water absorption capacity for breadmaking. Protein MWD parameters could be related to the association between DHV kernel level and water absorption. Specifically, flour protein fractions rich in gliadins and high‐molecular‐weight polymeric proteins in the SDS‐unextractable fraction were identified to have significant and positive correlations with both DHV kernels and flour water absorption levels. An example further showed the importance of flour water absorption on potential economic incentives that can be gained with having a greater percentage of vitreous kernels. This information could help the flour milling and baking industry to segregate the different subclasses of HRS wheat with varying DHV content for their intended end‐use applications.     

Relationship Between Solvent Retention Capacity and Protein Molecular Weight Distribution,Quality Characteristics,and Breadmaking Functionality of Hard Red Spring Wheat Flour

This research aims to investigate the relationship between the solvent retention capacity (SRC) test and quality assessment of hard red spring (HRS) wheat flour samples obtained from 10 HRS cultivars grown at six locations in North Dakota. The SRC values were significantly (P < 0.05) correlated with flour chemical components (protein, gluten, starch, and damaged starch contents, except arabinoxylan); with farinograph parameters (stability [FST], water absorption, peak time [FPT], and quality number); and with breadmaking parameters (baking water absorption [BWA], bread loaf volume [BLV], and symmetry). Differences in locations and cultivars contributed significantly to variation in quality parameters and SRC values. Suitability of SRC parameters for discriminatory analysis of HRS wheat flour is greatly influenced by molecular weight distribution (MWD) of SDS‐unextractable proteins. SRC parameters, except for sucrose SRC, showed significant (P < 0.01) and positive correlations with high‐molecular‐weight (HMW) polymeric proteins in SDS‐unextractable fractions, whereas only lactic acid SRC exhibited significant (P < 0.01) correlations with low‐molecular‐weight polymeric proteins. HMW polymeric proteins also exhibited positive associations with FPT, FST, BWA, and BLV. The discrepant variation in association of SRC parameters with respect to MWD of SDS‐unextractable proteins could improve segregation of HRS wheat flour samples for quality.     

Isolation and characterization of antimicrobial-resistant Escherichia coli from national horse racetracks and private horse-riding courses in Korea

Limited information is available regarding horse-associated antimicrobial resistant (AR) Escherichia (E.) coli. This study was designed to evaluate the frequency and characterize the pattern of AR E. coli from healthy horse-associated samples. A total of 143 E. coli (4.6%) were isolated from 3,078 samples collected from three national racetracks and 14 private horse-riding courses in Korea. Thirty of the E. coli isolates (21%) showed antimicrobial resistance to at least one antimicrobial agent, and four of the AR E. coli (13.3%) were defined as multi-drug resistance. Most of the AR E. coli harbored AR genes corresponding to their antimicrobial resistance phenotypes. Four of the AR E. coli carried class 1 integrase gene (intI1), a gene associated with multi-drug resistance. Pulsed-field gel electrophoretic analysis showed no genetic relatedness among AR E. coli isolated from different facilities; however, cross-transmissions between horses or horses and environments were detected in two facilities. Although cross-transmission of AR E. coli in horses and their environments was generally low, our study suggests a risk of transmission of AR bacteria between horses and humans. Further studies are needed to evaluate the risk of possible transmission of horse-associated AR bacteria to human communities through horse riders and horse-care workers.     

Hepatitis E virus as an emerging zoonotic pathogen

Hepatitis E outbreaks are a serious public health concern in developing countries. The disease causes acute infections, primarily in young adults. The mortality rate is approximately 2%; however, it can exceed 20% in pregnant women in some regions in India. The causative agent, hepatitis E virus (HEV), has been isolated from several animal species, including pigs. HEV genotypes 3 and 4 have been isolated from both humans and animals, and are recognized as zoonotic pathogens. Seroprevalence studies in animals and humans indirectly suggest that HEV infections occur worldwide. The virus is primarily transmitted to humans via undercooked animal meats in developed countries. Moreover, transfusion- and transplantation-mediated HEV infections have recently been reported. This review summarizes the general characteristics of hepatitis E, HEV infection status in animals and humans, the zoonotic transmission modes of HEV, and HEV vaccine development status.     

Apparent ileal digestibility of nutrients and amino acids in soybean meal,fish meal,spray‐dried plasma protein and fermented soybean meal to weaned pigs

This study sought to determine whether fermentation could increase apparent ileal digestibility (AID) of dry matter (DM), nitrogen (N), energy (E) and amino acids (AA) in fermented soybean meal (FSBM) greater than that of soybean meal (SBM) in weaned pigs. Four weaned pigs (10.00 ± 0.30 kg) were surgically equipped with T‐cannulas and randomly followed a 4 × 4 Latin square design of treatments (SBM, FSBM, fish meal and spray‐dried plasma protein). Overall, the fermentation process was able to reduce the amount of anti‐nutritional factors (ANF), including trypsin inhibitors, raffinose and stachyose, in the FSBM diet, which were significantly reduced by 39.4, 92.2, and 92.9%, respectively, as compared to the SBM diet. As a consequence of ANF reduction in FSBM, the AID of DM, N and E as well as AA was significantly greater with FSBM than SBM. Taken all together, the fermentation process improved the nutritional quality of SBM, due to ANF reduction, leading to improvement of digestibility of AA. As such, FSBM can be potentially used as a specialized feed ingredient, especially for young animal diets in an attempt to reduce diet costs.