Enterotoxigenicity and cytotoxicity of Bacillus thuringiensis strains and development of a process for Cry1Ac production

Bacillus thuringiensis is indistinguishable from Bacillus cereus except for the production of insecticidal crystal proteins (ICPs). B. thuringiensis strains may show enterotoxin profiles and toxin levels similar to those of B. cereus strains isolated from food-poisoning cases. It is important for the food industry and farmers to consider that with the application of B. thuringiensis strains to crops, their spores may be introduced into the human food chain. In this study, 59 B. thuringiensis strains were assayed for their hemolysin BL (HBL) using a BCET-RPLA kit and their cytotoxicity to Chinese hamster ovary (CHO) cells. The enterotoxin titer was as high as that of B. cereus diarrheal-type strain ATCC 49064. In an attempt to obtain a food safety strain for bioinsecticide use, in this study, a 3.5-kb cry1Ac DNA fragment was amplified with PCR from the total DNA of B. thuringiensis subsp. kurstaki CCRC 11502 and cloned into the Bacillus expression vector pHY300PLK. The alpha-amylase promoter, amyE, was then introduced into the promoter region and, afterward, the recombinant plasmid pHYe1Ac35 was introduced into a non-enterotoxigenic and non-cytotoxic B. thuringiensis subsp. kurstaki Tt14 strain. The transformant, without any detectable enterotoxigenicity or cytotoxicity, produced Cry1Ac toxin properly, and its insecticidal activity against Trichoplusia ni larvae was found to be satisfactory.     

Biochemical and molecular analyses to determine pyrethroid resistance in Aedes aegypti

Dengue fever is an important mosquito-borne viral disease in Taiwan. Insecticide resistance has been shown to significantly reduce the efficacy of vector control interventions. The detection of insecticide resistance is an important component in mosquito abatement programs. In this study, we used the insecticide-impregnated papers bioassay method to reveal high levels of resistance to permethrin in the LYPR and field strains of Aedes aegypti. We used the standard glass cylinder method to observe the knockdown effect of paralysis within 2 to 4 minutes after exposing mosquitoes to pyrethroid vapors. Biochemical assays showed elevated detoxification enzyme activities. Glutathione S-transferases, monooxygenases and β-esterases were the enzymes predominantly responsible for the permethrin resistance of Ae. aegypti in Taiwan. Molecular screening for common insecticide target-site mutations revealed the presence of V1023G and D1794Y mutations. Pearson’s correlation analysis was used to investigate the correlations between the allelic frequency of kdr mutation associated increase with the LC50 values of permethrin and the KT50 values of pyrethroid vaporizers. These findings will be used to assess resistance levels, estimate resistance potential, and formulate monitoring and resistance management strategies.     

Pulsed field gel electrophoresis for animal Salmonella enterica serovar Typhimurium isolates in Taiwan

Salmonella enterica subspecies enterica serovar Typhimurium is a common pathogen for humans and animals. In order to trace the clonal relationship and to find the circulating strains between human and animal isolates, chromosomal DNAs from 87 serovar Typhimurium strains isolated from animals (pigs were the majority) were subjected to XbaI and SpeI digestion and pulsed field gel electrophoresis (PFGE). For the 87 animal isolates, 38 PFGE pattern combinations were obtained. As the subtyping results from animal isolates were compared with those from the 45 human isolates, it was found that 14 of the animal isolates and 13 of the human isolates shared a common PFGE pattern combination, i.e., pattern XgSf (or called X5S4). When these human and animal isolates were subjected to antibiotic susceptibility test using 11 antibiotics, it was found that strains of pattern XgSf (X5S4) belong to a common antibiogram pattern which is tetracycline, gentamicin, ampicillin, streptomycin and chloramphenicol resistant. Since most of the animal and human strains in pattern XgSf were originally isolated from various areas over different years, strains of this PFGE pattern may be the most epidemic strains which circulating between human and animal sources.     

Inhibitory activity of natural occurring antioxidants on Thiyl radical-induced trans-arachidonic acid formation

trans-Fatty acids in humans not only may be obtained exogenously from food intake but also could be generated endogenously in tissues. The endogenous generation of trans-fatty acids, especially in the cell membranes induced by radical stress, is an inevitable source for the living species. Thiyl radicals generated from thiols act as the catalyst for the cis-trans isomerization of fatty acids. Arachidonic acid (5c,8c,11c,14c-20:4) with only two of the four double bonds deriving from linoleic acid in the diet can be used to differentiate the exogenous or endogenous formation of double bonds. The aim of this study is to evaluate the effective compounds in preventing thiyl radical-induced trans-arachidonic acid formation during UV irradiation in vitro. The trans-arachidonic acids were found to be 75% after 30 min UV irradiation of all-cis-arachidonic acid. Myricetin, luteolin, and quercetin had the highest thiyl radical scavenging activities, whereas sesamol, gallic acid, and vitamins A, C, and E had the lowest. The structures of flavonoids with higher thiyl radical scavenging activities were a 3',4'-o-dihydroxyl group in the B ring and a 2,3-double bond combined with a 4-keto group in the C ring. These effective compounds found in the present work may be used as lead compounds for the potential inhibitors in the formation of trans-fatty acids in vivo.     

Discharge estimation in lined irrigation canals by using surface velocity radar

This study proposes a surface velocity method for discharge estimation in lined irrigation canals. In this method, a surface velocity radar is used for surface velocity measurement. However, this radar cannot be used to obtain a point velocity on the water surface. A velocity profiler is used to measure the velocity distribution in the cross section of a canal. The measured data are used to obtain surface and mean velocities in verticals by applying a probabilistic velocity distribution equation. Subsequently, the linear relationship between the surface velocity measured by the surface velocity radar and the surface velocity estimated by the velocity distribution equation can be established. Additionally, the relationship between the estimated surface velocity and mean velocity in verticals can be obtained through linear regression; this relationship is a constant. Therefore, these two relationships can be used to derive a surface velocity coefficient for converting surface velocity to mean velocity in a vertical. The surface velocity coefficient used in lined irrigation canals is different from that used in natural streams. It is not constant and varies with the surface velocity observed by the radar. The feasibility of the proposed method is tested by conducting discharge estimation processes in five lined irrigation canals. The results show that the proposed method is reliable and accurate for estimating discharge in lined irrigation canals.     

Tailoring electrical transport across grain boundaries in polycrystalline graphene

Graphene produced by chemical vapor deposition (CVD) is polycrystalline, and scattering of charge carriers at grain boundaries (GBs) could degrade its performance relative to exfoliated, single-crystal graphene. However, the electrical properties of GBs have so far been addressed indirectly without simultaneous knowledge of their locations and structures. We present electrical measurements on individual GBs in CVD graphene first imaged by transmission electron microscopy. Unexpectedly, the electrical conductance improves by one order of magnitude for GBs with better interdomain connectivity. Our study suggests that polycrystalline graphene with good stitching may allow for uniformly high electrical performance rivaling that of exfoliated samples, which we demonstrate using optimized growth conditions and device geometry.     

Evaluation of suitable temperature range for post‐harvest processing of mud crabs through cardiac performance

Measurements of either heartbeats or heart rate variability provide important information on metabolic changes and stress responses. In this study, we aimed to determine the suitable temperature conditions for maintaining product quality for post‐harvest processing of mud crab (Scylla serrata) through measuring cardiac performance. A bundle of implanted electrodes was used to record the electrocardiogram, from which the heart rate (HR) and heart rate variability (HRV) were derived to evaluate the physiological performances of the crabs under tying at different seawater temperatures. The lowest standard deviation of HR was detected at the seawater temperature of 18°C for the group of ascending temperatures and at 10°C for the group of descending temperatures. Lower HRV was detected at the range from 10 to 14°C. The results suggest that the temperature range of 14–18°C is suitable to reduce variability in heart rate and may decrease physiological stress. These conditions might help maintain the quality of live crabs during post‐harvest processing and transportation.