Recent Developments in Enzymatic Antioxidant Defence Mechanism in Plants with Special Reference to Abiotic Stress.

The stationary life of plants has led to the evolution of a complex gridded antioxidant defence system constituting numerous enzymatic components, playing a crucial role in overcoming various stress conditions. Mainly, these plant enzymes are superoxide dismutase (SOD), catalase (CAT), peroxidase (POX), glutathione peroxidase (GPX), glutathione reductase (GR), glutathione S-transferases (GST), ascorbate peroxidase (APX), monodehydroascorbate reductase (MDHAR), and dehydroascorbate reductase (DHAR), which work as part of the antioxidant defence system. These enzymes together form a complex set of mechanisms to minimise, buffer, and scavenge the reactive oxygen species (ROS) efficiently. The present review is aimed at articulating the current understanding of each of these enzymatic components, with special attention on the role of each enzyme in response to the various environmental, especially abiotic stresses, their molecular characterisation, and reaction mechanisms. The role of the enzymatic defence system for plant health and development, their significance, and cross-talk mechanisms are discussed in detail. Additionally, the application of antioxidant enzymes in developing stress-tolerant transgenic plants are also discussed.

Effect of Soil Chemical Properties on the Occurrence and Distribution of Entomopathogenic Fungi in Portuguese Grapevine Fields.

Entomopathogenic fungi (EPF) contribute to different ecosystem services. However, factors affecting their natural occurrences in soil remain poorly understood. In a previous study, 81 soil samples were subjected to insect baiting using Galleria mellonella and Tenebrio molitor to isolate EPF from Portuguese vine farms. Here, soils yielding any of the four common EPF, i.e., Beauveria bassiana, Purpureocillium lilacinum, Metarhizium robertsii, and Clonostachys rosea f. rosea, were correlated with their chemical properties. Beauveria bassiana was negatively affected by higher available P (p = 0.02), exchangeable K-ions (p = 0.016) and positively affected by higher soil pH_H2O (p = 0.021). High exchangeable K-ions inhibited P. lilacinum (p = 0.011) and promoted C. rosea f. rosea (p = 0.03). Moreover, high available K also suppressed P. lilacinum (p = 0.027). Metarhizium robertsii was inhibited by higher organic matter content (p = 0.009), higher C:N (p = 0.017), total N (p = 0.007), and exchangeable Mg-ions (p = 0.026), and promoted by higher exchangeable Na-ions (p = 0.003). Nonetheless, mean comparisons and principal component analysis suggested that higher soil pH and exchangeable Ca-ions have contrasting effects on EPF occurrences, as they promote B. bassiana and inhibit M. robertsii. Herbicides did not seem to affect EPF presence. Overall, this study is among the first reports on the effects of soil chemistry on EPF other than Metarhizium, and will facilitate biological pest management approaches.

Advances in Entomopathogen Isolation: A Case of Bacteria and Fungi.

Entomopathogenic bacteria and fungi are quite frequently found in soils and insect cadavers. The first step in utilizing these microbes as biopesticides is to isolate them, and several culture media and insect baiting procedures have been tested in this direction. In this work, the authors review the current techniques that have been developed so far, in the last five decades, and display brief protocols which can be adopted for the isolations of these entomopathogens. Among bacteria, this review focuses on Serratia spp. and bacteria from the class Bacilli. Among fungi, the review focuses those from the order Hypocreales, for example, genera Beauveria, Clonostachys, Lecanicillium, Metarhizium, and Purpureocillium. The authors chose these groups of entomopathogenic bacteria and fungi based on their importance in the microbial biopesticide market.

First Demonstration of Clinical Fusarium Strains Causing Cross-Kingdom Infections from Humans to Plants.

Mycotoxins from the Fusarium genus are widely known to cause economic losses in crops, as well as high mortalities rates among immunocompromised humans. However, to date, no correlation has been established for the ability of Fusarium to cause cross-kingdom infection between plants and humans. The present investigation aims to fill this gap in the literature by examining cross-kingdom infection caused by Furasium strains isolated from non-immunocompromised or non-immunosuppressed humans, which were subsequently reinfected in plants and on human tissue. The findings document for the first time cross-kingdom infective events in Fusarium species, thus enhancing our existing knowledge of how mycopathogens continue to thrive in different hosts.

Fusarium, an Entomopathogen-A Myth or Reality?

The Fusarium species has diverse ecological functions ranging from saprophytes, endophytes, and animal and plant pathogens. Occasionally, they are isolated from dead and alive insects. However, research on fusaria-insect associations is very limited as fusaria are generalized as opportunistic insect-pathogens. Additionally, their phytopathogenicity raises concerns in their use as commercial biopesticides. Insect biocontrol potential of Fusarium is favored by their excellent soil survivability as saprophytes, and sometimes, insect-pathogenic strains do not exhibit phytopathogenicity. In addition, a small group of fusaria, those belonging to the Fusarium solani species complex, act as insect mutualists assisting in host growth and fecundity. In this review, we summarize mutualism and pathogenicity among fusaria and insects. Furthermore, we assert on Fusarium entomopathogenicity by analyzing previous studies clearly demonstrating their natural insect-pathogenicity in fields, and their presence in soils. We also review the presence and/or production of a well-known insecticidal metabolite beauvericin by different Fusarium species. Lastly, some proof-of-concept studies are also summarized, which demonstrate the histological as well as immunological changes that a larva undergoes during Fusarium oxysporum pathogenesis. These reports highlight the insecticidal properties of some Fusarium spp., and emphasize the need of robust techniques, which can distinguish phytopathogenic, mutualistic and entomopathogenic fusaria.

Soil Chemical Properties Barely Perturb the Abundance of Entomopathogenic Fusarium oxysporum: A Case Study Using a Generalized Linear Mixed Model for Microbial Pathogen Occurrence Count Data.

Fusarium oxysporum exhibits insect pathogenicity-however, generalized concerns of releasing phytopathogens within agroecosystems marred its entomopathogenicity-related investigations. In a previous study, soils were sampled from Douro vineyards and adjacent hedgerows. In this study, 80 of those soils were analyzed for their chemical properties and were subsequently co-related with the abundance of entomopathogenic F. oxysporum, after insect baiting of soils with Galleria mellonella and Tenebrio molitor larvae. The soil chemical properties studied were organic matter content; total organic carbon; total nitrogen; available potassium; available phosphorus; exchangeable cations, such as K⁺, Na⁺, Ca2+, and Mg2+; pH; total acidity; degree of base saturation; and effective cation exchange capacity. Entomopathogenic F. oxysporum was found in 48 soils, i.e., 60% ± 5.47%, of the total soil samples. Out of the 1280 insect larvae used, 93, i.e., 7.26% ± 0.72%, were found dead by entomopathogenic F. oxysporum. Stepwise deletion of non-significant variables using a generalized linear model was followed by a generalized linear mixed model (GLMM). A higher C:N (logarithmized) (p < 0.001) and lower exchangeable K⁺ (logarithmized) (p = 0.008) were found significant for higher fungal abundance. Overall, this study suggests that entomopathogenic F. oxysporum is robust with regard to agricultural changes, and GLMM is a useful statistical tool for count data in ecology.

Entomopathogenic fungi in Portuguese vineyards soils: suggesting a 'Galleria-Tenebrio-bait method' as bait-insects Galleria and Tenebrio significantly underestimate the respective recoveries of Metarhizium (robertsii) and Beauveria (bassiana).

Entomopathogenic fungi (EPF) are the natural enemies of insect-pests. However, EPF recoveries can be influenced by the soil habitat-type(s) incorporated and/or the bait-insect(s) used. Galleriamellonella (GM) as bait-insect, i.e. 'Galleria-bait', is arguably the most common methodology, which is sometimes used solely, to isolate EPF from soils. Insect baiting using Tenebriomolitor (TM) has also been employed occasionally. Here 183 soils were used to estimate the functional diversity of EPF in Portuguese Douro vineyards (cultivated habitat) and adjacent hedgerows (semi-natural habitat), using the TM bait method. Moreover, to study the effect of insect baiting on EPF recovery, 81 of these 183 soil samples were also tested for EPF occurrences using the GM bait method. Twelve species were found in 44.26% ± 3.67% of the total of 183 soils. Clonostachysroseaf.rosea was found in maximum soils (30.05% ± 3.38%), followed by Beauveriabassiana (12.57% ± 2.37%), Purpureocilliumlilacinum (9.29% ± 2.14%) and Metarhiziumrobertsii (6.01% ± 1.75%). Beauveriapseudobassiana (P < 0.001), C.roseaf.rosea (P = 0.006) and Cordycepscicadae (P=0.023) were isolated significantly more from hedgerows, highlighting their sensitivities towards agricultural disturbances. Beauveriabassiana (P = 0.038) and M.robertsii (P = 0.003) were isolated significantly more using GM and TM, respectively. Principal component analysis revealed that M.robertsii was associated both with TM baiting and cultivated habitats, however, B.bassiana was slightly linked with GM baiting only. Ecological profiles of B.bassiana and P.lilacinum were quite similar while M.robertsii and C.roseaf.rosea were relatively distant and distinct. To us, this is the first report on (a) C.cicadae isolation from Mediterranean soils, (b) Purpureocilliumlavendulum as an EPF worldwide; and (c) significant recoveries of M.robertsii using TM over GM. Overall, a 'Galleria-Tenebrio-bait method' is advocated to study the functional diversity of EPF in agroecosystems.

Multigene assessment of the species boundaries and sexual status of the basidiomycetous yeasts Cryptococcus flavescens and C. terrestris (Tremellales).

Cryptococcus flavescens and C. terrestris are phenotypically indistinguishable sister species that belong to the order Tremellales (Tremellomycetes, Basidiomycota) and which may be mistaken for C. laurentii based on phenotype. Phylogenetic separation between C. flavescens and C. terrestris was based on rDNA sequence analyses, but very little is known on their intraspecific genetic variability or propensity for sexual reproduction. We studied 59 strains from different substrates and geographic locations, and used a multilocus sequencing (MLS) approach complemented with the sequencing of mating type (MAT) genes to assess genetic variation and reexamine the boundaries of the two species, as well as their sexual status. The following five loci were chosen for MLS: the rDNA ITS-LSU region, the rDNA IGS1 spacer, and fragments of the genes encoding the largest subunit of RNA polymerase II (RPB1), the translation elongation factor 1 alpha (TEF1) and the p21-activated protein kinase (STE20). Phylogenetic network analyses confirmed the genetic separation of the two species and revealed two additional cryptic species, for which the names Cryptococcus baii and C. ruineniae are proposed. Further analyses of the data revealed a high degree of genetic heterogeneity within C. flavescens as well as evidence for recombination between lineages detected for this species. Strains of C. terrestris displayed higher levels of similarity in all analysed genes and appear to make up a single recombining group. The two MAT genes (STE3 and SXI1/SXI2) sequenced for C. flavescens strains confirmed the potential for sexual reproduction and suggest the presence of a tetrapolar mating system with a biallelic pheromone/receptor locus and a multiallelic HD locus. In C. terrestris we could only sequence STE3, which revealed a biallelic P/R locus. In spite of the strong evidence for sexual recombination in the two species, attempts at mating compatible strains of both species on culture media were unsuccessful.

Cross-species gene expression analysis of species specific differences in the preclinical assessment of pharmaceutical compounds.

Animals are frequently used as model systems for determination of safety and efficacy in pharmaceutical research and development. However, significant quantitative and qualitative differences exist between humans and the animal models used in research. This is as a result of genetic variation between human and the laboratory animal. Therefore the development of a system that would allow the assessment of all molecular differences between species after drug exposure would have a significant impact on drug evaluation for toxicity and efficacy. Here we describe a cross-species microarray methodology that identifies and selects orthologous probes after cross-species sequence comparison to develop an orthologous cross-species gene expression analysis tool. The assumptions made by the use of this orthologous gene expression strategy for cross-species extrapolation is that; conserved changes in gene expression equate to conserved pharmacodynamic endpoints. This assumption is supported by the fact that evolution and selection have maintained the structure and function of many biochemical pathways over time, resulting in the conservation of many important processes. We demonstrate this cross-species methodology by investigating species specific differences of the peroxisome proliferator-activator receptor (PPAR) α response in rat and human.

Glass capillary microfluidics for production of monodispersed poly (DL-lactic acid) and polycaprolactone microparticles: experiments and numerical simulations.

HYPOTHESIS: Droplet size in microfluidic devices is affected by wettability of the microfluidic channels. Three-dimensional countercurrent flow focusing using assemblies of chemically inert glass capillaries is expected to minimize wetting of the channel walls by the organic solvent. EXPERIMENTS: Monodispersed polycaprolactone and poly(lactic acid) particles with a diameter of 18-150 µm were produced by evaporation of solvent (dichloromethane or 1:2 mixture of chloroform and toluene) from oil-in-water or water-in-oil-in-water emulsions produced in three-dimensional flow focusing glass capillary devices. The drop generation behaviour was simulated numerically using the volume of fluid method. FINDINGS: The numerical results showed good agreement with high-speed video recordings. Monodispersed droplets were produced in the dripping regime when the ratio of the continuous phase flow rate to dispersed phase flow rate was 5-20 and the Weber number of the dispersed phase was less than 0.01. The porosity of polycaprolactone particles increased from 8 to 62% when 30 wt% of the water phase was incorporated in the organic phase prior to emulsification. The inner water phase was loaded with 0.156 wt% lidocaine hydrochloride to achieve a sustained drug release. 26% of lidocaine was released after 1 h and more than 93% of the drug was released after 130 h.