Detection of the Defoliating Pathotype of Verticillium dahliae in Infected Olive Plants by Nested PCR

Spread of Verticillium wilt into newly established olive orchards in Andalucía, southern Spain, has caused concern in the olive industry in the region. This spread may result from use of Verticillium dahliae-infected planting material, which can extend distribution of the highly virulent, defoliating (D) pathotype of V. dahliae to new areas. In this study, a molecular diagnostic method for the early in planta detection of D V. dahliae was developed, aimed especially at nursery-produced olive plants. For this purpose, new primers for nested PCR were designed by sequencing a 992-bp RAPD marker of the D pathotype. The use of the specific primers and different nested-PCR protocols allowed the detection of V. dahliae pathotype D DNA in infected root and stem tissues of young olive plants. Detection of the pathogen was effective from the very earliest moments following inoculation of olive plants with a V. dahliae pathotype D conidia suspension as well as in inoculated, though symptomless, plants.     

Induction of an antioxidant enzyme system and other oxidative stress markers associated with compatible and incompatible interactions between chickpea (Cicer arietinum L.) and Fusarium oxysporum f. sp.ciceris

To ascertain if active oxygen species play a role in fusarium wilt of chickpea caused by Fusarium oxysporum f. sp. ciceris, the degree of lipid peroxidation (malondialdehyde formation) and the activity levels of diamine oxidase (DAO), an apoplastic H₂O₂-forming oxidase, and several antioxidant enzymes, namely ascorbate peroxidase (APX), catalase (CAT), glutathione reductase (GR), guaiacol-dependent peroxidase (GPX) and superoxide dismutase (SOD), were determined spectrophotometrically in roots and stems of ‘WR315’ (resistant) and ‘JG62’ (susceptible) chickpea cultivars inoculated with the highly virulent race 5 of the pathogen. Moreover, APX, CAT, GPX and SOD were also analysed in roots and stems by gel electrophoresis and activity staining; and the protein levels of APX and SOD in roots were determined by Western blotting. In roots, infection by the pathogen increased lipid peroxidation and CAT and SOD activities, although such responses occurred earlier in the incompatible compared with the compatible interactions. APX, GPX and GR activities were also increased in infected roots, but only in the compatible interaction. In stems, infection by the pathogen increased lipid peroxidation and APX, CAT, SOD and GPX activities only in the compatible interaction, and DAO activity only in the incompatible one. In general, electrophoregrams agreed with the activity levels determined spectrophotometrically and did not reveal any differences in isoenzyme patterns between cultivars or between infected and non-infected plants. Further, Western blots revealed an increase in the root protein levels of APX in the compatible interaction and in those of SOD in both compatible and incompatible interactions. In conclusion, whereas enhanced DAO activity in stems, and earlier increases in lipid peroxidation and CAT and SOD activities in roots, can be associated with resistance to fusarium wilt in chickpea, the induction of the latter three parameters in roots and stems along with that of APX, GR (only in roots) and GPX (only in stems) activities are rather more associated with the establishment of the compatible interaction.     

Effect of chitin on biological control activity of Bacillus spp. and Trichoderma harzianum against root rot disease in pepper (Capsicum annuum) plants

Two bacterial isolates and one strain of Trichoderma harzianum were tested alone and in combination with chitin for efficacy in control of root rot disease caused by Phytophthora capsici and Rhizoctonia solani in pepper plants under greenhouse conditions. These bacteria (Bacillus subtilis HS93 and B. licheniformis LS674) were isolated from repeatedly washed roots of pepper plants. In in vitro assays, HS93, LS674 and T. harzianum were antagonistic against P. capsici and R. solani and produced high levels of chitinase. Seed treatment and root drenching with bacterial suspensions of HS93 with 0.5% chitin was more effective against Phytophthora and Rhizoctonia root rot than addition of the organisms without chitin. LS674 and T. harzianum reduced Rhizoctonia but not Phytophthora root rot. In two greenhouse tests, seed treatment and root drenching with HS93 amended with chitin enhanced its biocontrol activity against P. capsici but not on R. solani. The effects of LS674 and T. harzianum against R. solani were significantly enhanced when they were used as suspensions with 0.5% chitin for root drenching, but this had no effect on P. capsici. In both greenhouse experiments, the use of 0.5% chitin alone for root drenching reduced Rhizoctonia root rot. Reduction of root rot disease was accompanied by increased yield. These results show that the antagonistic activity of HS93, LS674 and T. harzianum may be stimulated by chitin resulting in significant improvements in their effectiveness against pathogens.     

Distribution of alpha‐neoendorphin,ACTH (18–39) and beta‐endorphin (1–27) in the alpaca brainstem

Using an immunocytochemical technique, we have studied in the alpaca brainstem the distribution of immunoreactive structures containing prodynorphin (alpha‐neoendorphin)‐ and pro‐opiomelanocortin (adrenocorticotrophin hormone (18–39) (ACTH), beta‐endorphin (1–27))‐derived peptides. No peptidergic‐immunoreactive cell body was observed. Immunoreactive fibres were widely distributed, although in most of the brainstem nuclei the density of the peptidergic fibres was low or very low. In general, the distribution of the immunoreactive fibres containing the peptides studied was very similar. A close anatomical relationship occurred among the fibres containing alpha‐neoendorphin, ACTH or beta‐endorphin (1–27), suggesting a functional interaction among the three peptides in many of the brainstem nuclei. The number of fibres belonging to the prodynorphin system was higher than that of the pro‐opiomelanocortin system. A moderate/low density of immunoreactive fibres was observed in 65.11% (for alpha‐neoendorphin (1–27)), 18.18% (for ACTH) and 13.95% (for beta‐endorphin) of the brainstem nuclei/tracts. In the alpaca brainstem, a high density of immunoreactive fibres was not observed. The neuroanatomical distribution of the immunoreactive fibres suggests that the peptides studied are involved in auditory, motor, gastric, feeding, vigilance, stress, respiratory and cardiovascular mechanisms, taste response, sleep‐waking cycle and the control of pain transmission.     

Digestive Enzyme Activity during Larval Development of Black Snook,Centropomus nigrescens

Black snook, Centropomus nigrescens, have been identified as a promising candidate for aquaculture although, like many of the Centropomid species, high mortality associated with early larval stages presents a significant bottleneck to their commercialization. The digestive capacity of black snook larvae throughout the first 37 d after hatch (d.a.h.) was evaluated by quantifying digestive enzyme activities using biochemical techniques. Results showed that black snook larvae have alkaline proteases at hatching, which are known to be important during the first days of feeding for digestion. Toward the end of the study, acid proteases concentration increased (37 d.a.h.). Enzymes for lipid digestion, pancreatic lipase and bile salt‐activated lipase, were already present in the larvae before exogenous feeding commenced, and their activity increased with age and growth (length). Intracellular digestion, measured as the activity of leucine‐alanine peptidase, was high early on (5 d.a.h.) and decreased as development progressed (next 32 d). In contrast, alkaline phosphatase activity was lowest at first feeding and subsequently increased with age. Overall patterns in enzyme activity suggest the possibility of live feed weaning before 32 d.a.h. if artificial diets can be properly balanced.     

How does water and salt stress affect the germination and initial growth of Brazilian soya bean cultivars?

The effects of water and salt stress on rate of germination and seedling growth were investigated under laboratory conditions in 46 soya bean genotypes from Central-West region of Brazil to verify how these stresses may limit crop establishment during the initial growth stage and also to identify the most tolerant genotypes to drought and salinity. Mild water and salt stresses were imposed by seed exposure to –0.20 MPa iso-osmotic solutions with polyethylene glycol—PEG 6000 (119.57 g/L) or NaCl (2.357 g/L) for 12 days at 25°C. The germination percentage, seedling length and seedling dry matter were measured, and then, salt or drought tolerance indexes were calculated. The “NS 5909 RG,” “NS 7000 IPRO,” “NS 7338IPRO,” “FPS Solimões RR,” “NS 5151 IPRO,” “SYN 13610 IPRO,” “LG 60177 IPRO,” “NS 6909 IPRO” and “BMX Desafio RR” were identified as the most drought-tolerant genotypes, whereas under salinity conditions, the genotypes “5D 615 RR,” “BMX Desafio RR,” “5D 6215 IPRO” and “BMX Ponta IPRO” were identified as tolerant. The “BMX Desafio RR” is the genotype most adapted to both stress conditions and, therefore, should be used under conditions of water shortage and excess salt in the soil at sowing time.     

Histopathological findings in farmed rainbow trout (Oncorhynchus mykiss) naturally infected with 3 different Aeromonas species

This study describes the macroscopic and microscopic lesions in farmed rainbow trout (Oncorhynchus mykiss) naturally infected with genetically identified Aeromonas salmonicida, A. hydrophila, and A. veronii species. The genus Aeromonas includes bacteria that naturally inhabit both waterways and organisms. At least 27 Aeromonas species have been identified to date, some of which can cause significant economic losses in aquaculture. As up to 68.8% of Aeromonas isolates may be misidentified in routine biochemical and phenotypic tests, however, reported cases of Aeromonas infection in fish may be wrongly identified. Our findings confirmed that the 3 Aeromonas species studied are associated with septicemia and dermal lesions in rainbow trout.     

SALTIRSOIL: a simulation model for the mid to long‐term prediction of soil salinity in irrigated agriculture

The SALTIRSOIL model predicts soil salinity, sodicity and alkalinity in irrigated land using basic information on soil, climate, crop, irrigation management and water quality. It extends the concept of the WATSUIT model to include irrigation and crop management practices, advances in the calculation of evapotranspiration and new algorithms for the water stress coefficient and calculation of electrical conductivity. SALTIRSOIL calculates the soil water balance and soil solution concentration over the year. A second module, SALSOLCHEM, calculates the inorganic ion composition of the soil solution at equilibrium with soil calcite and gypsum at the soil’s CO₂ partial pressure. Results from comparing predicted and experimentally determined concentrations, observations and predictions of pH, alkalinity and calcium concentration in calcite‐saturated solutions agree to the second significant figure; in gypsum‐saturated solutions the standard difference between observations and predictions is <3% in absolute values. The algorithms in SALTIRSOIL have been verified and SALSOLCHEM validated for the reliable calculation of soil salinity, sodicity and alkalinity at water saturation in well‐drained irrigated lands. In simulations for horticultural crops in southeast Spain, soil solution concentration factors at water saturation, quotients of electrical conductivity (EC₂₅) at saturation to electrical conductivity in the irrigation water, and quotients of sodium adsorption ratio (SAR) are very similar to average measured values for the area.