Challenges facing arthropod biological control: identifying traits for genetic improvement of predators in protected crops

Biological control is an efficient pest control method but there are still limitations that are hindering its wider adoption. Genetic improvement of biological control agents (BCAs) can help to overcome these constraints, but the choice of key attributes for better performance that need to be selected is still an open question. Several characteristics have been suggested but the harsh reality is that selective breeding of BCAs has received a lot of attention but resulted in very little progress. Identifying the appropriate traits to be prioritized may be the first step to reverse this situation. In our opinion, the best way is to look at the factors limiting the performance of key BCAs, especially generalist predators (pesticide compatibility, prey‐density dependence, non‐suitable crops, and extreme environmental conditions), and according to these challenges, to choose the attributes that would allow BCAs to overcome those limitations. The benefits of selection for higher resistance to toxins, whether artificially applied (pesticides) or plant produced (plant defenses); increased fitness when feeding on non‐prey food (supplemented or plant‐derived); and better adaptation to extreme temperature and humidity are discussed. In conclusion, genetic improvement of BCAs can bring about new opportunities to biocontrol industry and users to enhance biocontrol resilience. © 2020 Society of Chemical Industry     

Simultaneous and sensitive detection of three foodborne pathogens by multiplex PCR, capillary gel electrophoresis, and laser-induced fluorescence

The simultaneous detection of Staphylococcus aureus, Listeria monocytogenes, and Salmonella spp. has been approached by a new multiplex PCR-based procedure followed by capillary gel electrophoresis with laser-induced fluorescence detection (multiplex-PCR-CGE-LIF). As compared to slab gel electrophoresis, the use of CGE-LIF improved from 10- to 1000-fold the sensitivity of the multiplex PCR analysis, allowing the detection of 2.6 x 10(3) cfu mL(-1) of S. aureus, 570 cfu mL(-1) of L. monocytogenes, and 790 cfu mL(-1) of Salmonella in artificially inoculated food, without enrichment. Following 6 h of enrichment, as low as 260, 79, and 57 cfu mL(-1) of S. aureus, L. monocytogenes, and Salmonella, respectively, were detected. The CGE-LIF method is shown to be reproducible, providing relative standard deviation (RSD) values lower than 0.8% for analysis time and lower than 5.8% for peak areas. The multiplex-PCR-CGE-LIF proved a powerful analytical tool to detect various food pathogens simultaneously in a fast, reproducible, and sensitive way.     

Beta-carotene isomer composition of sub- and supercritical carbon dioxide extracts. Antioxidant activity measurement

In the present work sub- and supercritical extraction conditions using carbon dioxide were studied in order to obtain extracts with different compositions from the green microalgae Dunaliella salina. Different compositions of beta-carotene isomers were identified in the extracts by using HPLC-DAD. Also, antioxidant activity of the extracts was measured using a TEAC assay. An experimental design was applied considering two factors, extraction pressure and temperature, in a wide range of values, trying to maximize the extraction yield. Higher yields were obtained at high pressures and low temperatures, that is, at higher CO2 densities. Attempts were made to correlate the antioxidant activity of the extracts with their chemical composition by means of principal component analysis. A certain relationship was found between their antioxidant activity and the isomeric composition of beta-carotenes. As a result, an original equation is proposed to predict the antioxidant activity of extracts from D. salina in terms of the ratio 9-cis-beta-carotene/all-trans-beta-carotene, the concentration of alpha-carotene, and, especially, the concentration of 9-cis-beta-carotene.     

Optimization of the extraction of antioxidants from Dunaliella salina microalga by pressurized liquids

In this work, extraction of antioxidant compounds from Dunaliella salina microalga is optimized by combining pressurized liquid extraction (PLE) and experimental design (three-level factorial design) with three different solvents (hexane, ethanol, and water). Two main factors were considered, the extraction temperature (40, 100, and 160 degrees C) and the extraction time (5, 17.5, and 30 min). As response variables, the extraction yield (percent dry weight/initial weight) and the antioxidant activity of the extracts (determined using the TEAC method) were used. The parameters of the model were estimated by multiple linear regression. Results showed that the extraction temperature was the factor having the strongest influence (positive) on the two response variables. The best yields were obtained with ethanol at the higher extraction temperature and time tested. Besides, although hexane extracts provided the best antioxidant activity, ethanol extracts were also very active. The chemical characterization of ethanol extracts was carried out using HPLC-DAD, and attempts have been made to correlate their chemical composition with the antioxidant activity measured. Results pointed out that the extracts contained, besides all-trans-beta-carotene and isomers, several different minor carotenoids that seemed to make a contribution to the antioxidant activity of the extracts.     

Ultrasensitive detection of genetically modified maize DNA by capillary gel electrophoresis with laser-induced fluorescence using different fluorescent intercalating dyes

In this work, four different fluorescent intercalating dyes are compared for the ultrasensitive CGE-LIF detection of DNA from transgenic maize in flours. The fluorescent intercalating dyes compared are YOPRO-1, SYBR-Green-I, Ethidium bromide (EthBr), and EnhanCE. For all the four dyes optimum concentrations are established, and efficient separations of DNA fragments ranging in size from 80 to 1000 bp are obtained. The comparative study demonstrates that SYBR-Green-I and YOPRO-1 provide better limits of detection (LODs) than EnhanCE or EthBr (i.e., LODs are, respectively, 700, 1000, 11300, and 97400 zmol, calculated for a 200-bp DNA fragment). Separations using YOPRO-1 are faster than those using SYBR-Green-I (30 min vs 47 min for the analysis of the 80-1000 bp DNA fragments). Also, separations using YOPRO-1 are more efficient than those using SYBR-Green-I (e.g., 2.4 x 10(6) plates/m vs 1.6 x 10(6) plates/m, respectively, calculated for the 200-bp fragment). Also, buffer depletion and cost per analysis are worse with SYBR-Green-I than with YOPRO-1. Therefore, YOPRO-1 was selected as the preferred intercalating dye. Using this fluorescent compound, analysis time reproducibility for the CGE-LIF separation of the DNA fragments is determined to be better than 1.7% (% RSD, n = 10) within the same day, and better than 1.9% (% RSD, n = 30) for three different days. Moreover, the fluorescence signal obtained using this dye is shown to vary linearly with the DNA concentration in the range studied, i.e., 1-500 ng/microL. It is demonstrated that by using this method 0.01% of transgenic maize can be detected in flour by direct injection of the PCR-amplified sample.     

Characterization of protein fractions from Bt-transgenic and non-transgenic maize varieties using perfusion and monolithic RP-HPLC. Maize differentiation by multivariate analysis

Protein fractions from transgenic Bt and non-transgenic maize varieties, extracted by the Osborne solvent fraction procedure, were characterized for the first time by perfusion and monolithic RP-HPLC in very short analysis times. Albumins and globulins from different transgenic Bt maizes as well as from their non-transgenic isogenic varieties were eluted in four peaks using perfusion RP-HPLC, whereas prolamins and glutelins were separated in seven peaks. Monolithic RP-HPLC enabled the separation of maize proteins in a large number of peaks showing 6 and 10 main peaks for albumins and globulins, respectively. Prolamins migrated at retention times higher than 5 min as seven peaks, whereas glutelins were separated in three main peaks appearing at retention times higher than 6.0 min. Moreover, chromatograms of the whole protein extracts showed 8 and 11 components for perfusion and monolithic RP-HPLC, respectively. A comparison of the chromatograms of the whole protein extracts relative to transgenic and non-transgenic varieties evidenced quantitative differences on the percentages of area, mainly for peaks 2 and 3 by perfusion RP-HPLC and for peaks 3 and 7 by monolithic RP-HPLC. A discriminant analysis based on these proteic profiles was carried out to classify and predict transgenic Bt maize lines, achieving 100% correct classification using perfusion RP-HPLC.     

Comparison of Spore Inoculated and Vegetative Propagated Cultivation Methods of <Emphasis Type="Italic">Gracilaria chilensis</Emphasis> in an Integrated Seaweed and Fish Cage Culture

In Chile the integration of Gracilaria chilensis with salmon culture has shown high potential. Seaweed integrated aquaculture is of great interest as it allows waste recycling within fed cage aquaculture. The development of economically feasible suspended methods of seaweed cultivation is therefore of high importance. Hence, production and performance of two suspended Gracilaria cultivation methods, spore inoculated ropes and ropes with twined field collected seaweed, were studied in open water. The production from spore-seeded ropes was comparable to that of twined ropes for the first month of culture. Thereafter, the twined ropes had a significantly higher productivity. Fish farm wastes had no significant fertilizing effect upon Gracilaria growth rate. In addition, spore-originated thalli and field collected thalli were compared under laboratory conditions and in suspended culture using the same cultivation method. Spore-originated thalli had a 50% lower growth rate than the field collected thalli under laboratory conditions; however, no significant differences were detected in the field. Also, the occurrence of spore coalescence growth enhancement was not significant on the spore-seeded ropes. It was concluded that spore-originated cultivation techniques could be of interest for an integrated open seawater aquaculture system due to the high levels of Gracilaria polymorphism. This would result in greater adaptability to environmental variations, and a continuous supply of restocking material.     

Metabolite Profiling of Rosemary Cell Lines with Antiproliferative Potential against Human HT-29 Colon Cancer Cells

Plant Foods for Human Nutrition - Rosemary (Rosmarinus officinalis) is a culinary and medicinal plant used in food and pharmaceutical industry. The wide range of biological activities is mainly...     

Modeling the Interfacial Interactions between CrtS and CrtR from Xanthophyllomyces dendrorhous , a P450 System Involved in Astaxanthin Production

Xanthophyllomyces dendrorhous is a natural source of astaxanthin, a carotenoid widely used in the food industry. In this yeast, astaxanthin is synthesized from β-carotene by a cytochrome P450, CrtS, which depends on CrtR, the four-domain cytochrome P450 reductase (CPR). Although Saccharomyces cerevisiae has an endogenous CPR (ScCPR), expression of CrtS does not result in astaxanthin production unless it is coexpressed with CrtR. Assuming that CrtS could interact with the FMN-binding domain of either CrtR or ScCPR (XdFMNbd and ScFMNbd, respectively), the aim of this work was to identify possible interaction differences between these alternative complexes by protein modeling and short molecular dynamics simulations. Considering the recently proposed membrane orientation of a mammalian P450, our CrtS-CrtR model predicts that both N-terminal ends stand adjacent to the membrane plane, allowing their anchoring. Compared with the possible interface between CrtS and both FMNbd, the Xanthophyllomyces system appears to be stabilized by more saline bridges.