Effect of Pichia membranaefaciens in combination with salicylic acid on postharvest blue and green mold decay in citrus fruits

The separate or combined effects of Pichia membranaefaciens and salicylic acid (SA) on the control of blue and green mold decay in citrus fruits were investigated. Results indicate that combining P. membranaefaciens (1 × 10⁸ CFU ml⁻¹) with SA (10 μg ml⁻¹) either in a point-inoculated or dipped treatment provided a more effective control of blue and green mold than separately applying yeast or SA. SA (10 μg ml⁻¹) did not significantly affect P. membranaefaciens growth in vitro but slightly increased the yeast population in fruit wounds. P. membranaefaciens plus SA effectively enhanced the phenylalanine ammonia-lyase, peroxidase, polyphenoloxidase, chitinase, and β-1,3-glucanase activities and stimulated the synthesis of phenolic compounds. The combined treatment did not impair quality parameters such as weight loss or titratable acidity, but resulted in low average natural infection incidence and increased total soluble solids and ascorbic acid contents in citrus fruits after 14 d at 20 °C.     

Effect of preharvest spraying Cryptococcus laurentii on postharvest decay and quality of strawberry

Application of a suspension (log 8.0/ml) of Cryptococcus laurentii prior to harvest led to a reduction in Botrytis cinerea decay of strawberries stored at 4 or 20 °C, for 12 or 4 days, respectively. The frequency of spraying antagonist significantly influenced disease incidence in strawberry fruit. The best inhibition of disease was achieved when fruit sprayed C. laurentii with three applications that began 6 days prior to harvest, and the incidence of gray mold and natural decay treated with this method was 21% and 11%, compared with 88% and 62% in the control after storage at 20 °C for 4 days. A similar result occurred in the treated fruit after storage at 4 °C for 12 days. Dilation plate counts on Rose Bengal agar and scanning electron microscopy results showed that three applications with C. laurentii at 6, 3 and 0 days before harvest improved its ability to colonize the epidermis of strawberry fruit in the greenhouse and during storage compared to single application. In addition, antagonistic yeast spraying before harvest could reduce the weight loss, delay the decrease of the firmness and ascorbic acid, but had no significant effect on the contents of soluble solids, titratable acidity and fruit surface color. These results suggested that preharvest spraying with C. laurentii might be a promising alternative to fungicide application for decay control of strawberry.     

Efficacy of the antagonist Aureobasidium pullulans PL5 against postharvest pathogens of peach,apple and plum and its modes of action

The efficacy of Aureobasidium pullulans PL5 against different postharvest pathogens of fruits (Monilinia laxa on plums and peaches, Botrytis cinerea and Penicillium expansum on apples) were evaluated under storage conditions when applied at 10⁸ cells ml⁻¹ and their interactions were studied in vitro and in vivo to discover the possible modes of action. Under 1.2 °C and 95% relative humidity (RH) for 28 days, the efficacy of PL5 against M. laxa on plums was 45%, reducing disease incidence from 78% to 43%. Under 1 °C and 95% RH for 21 days, the efficacy against M. laxa on peaches was 63%, reducing disease incidence from 79% to 29%. Under 4 °C and 95% RH for 45 days, the efficacy against B. cinerea and P. expansum on apples was 56% and 46%, respectively. In Lilly–Barnett minimal salt medium with the fungal cell walls of pathogens as sole carbon source, PL5 produced β-1,3-glucanase, exo-chitinase and endo-chitinase. Nutrient concentrations had significant effect on pathogen growth reduction by PL5. No attachment was observed in antagonist–pathogen interactions in vitro or in vivo. PL5 completely inhibited pathogen spore germination in PDB at 10⁸ cells ml⁻¹, whereas at 10⁶ cells ml⁻¹ the efficacy was significantly decreased. However, inactivated cells and culture filtrate of PL5 had no effect on pathogen spore germination and germ tube elongation. Our results showed that A. pullulans PL5 could be introduced in commercial formulations to control postharvest pathogens on fruits and its activity was based on secretion of lytic enzymes and competition for nutrients.     

Postharvest biocontrol of Monilinia laxa,Monilinia fructicola and Monilinia fructigena on stone fruit by two Aureobasidium pullulans strains

The antagonistic effects of yeasts, L1 and L8, isolated from carposphere of ‘Redhaven’ peaches were tested for the first time in the same experiment against three Monilinia species (Monilinia laxa, Monilinia fructicola and Monilinia fructigena) in in vitro and in vivo trials. The two antagonists were selected after preliminary assays for their ability to reduce brown rot in peaches and nectarines, and both were identified by molecular and morphological tools as Aureobasidium pullulans. In in vivo trials, neither the autoclaved cells, nor the sterile culture filtrates of either antagonist showed any significant reduction of rot incidence produced by inocula of the three Monilinia species, while the washed cells of L1 and L8 completely inhibited M. laxa and M. fructicola rots and reduced M. fructigena infections by 70% and 90%, respectively. In other trials, nectarines treated with antagonist cells and inoculated with the pathogens were stored at 0 °C for 21 days, plus 7 days at 20 °C. The low temperature reduced brown rot development, since all fruit were free from disease symptoms on removal from cold storage. However after 7 d at 20 °C, untreated fruit were rotted over 45% depending on the Monilinia species but the antagonists completely inhibited M. laxa and M. fructicola, while M. fructigena infections were reduced by 89.8% and 91.2% by L1 and L8, respectively. For both strains, 10⁸ CFU ml^?1 was the most active concentration, although L1 showed good activity at a concentration of 10⁷ CFU ml^?1. Isolate L8 at the concentration of 10⁷ CFU ml^?1 was ineffective against M. fructicola and M. fructigena, showing no difference between treated fruit and the control, excepting the case of nectarines inoculated with M. laxa, where L8 at the concentration of 10⁷ CFU ml^?1 reduced the brown rot infections with respect to the control. The increase in population density of A. pullulans strains L1 and L8 in the wounds of nectarines stored at 0° or 20 °C was low but sufficient to control brown rot. In conclusion, the present preliminary study identified two antagonistic strains of A. pullulans as active ingredients for the development of biofungicides for postharvest application against three Monilinia species that are responsible for high economic losses in stone fruit crops.     

Stability of the intra- and extracellular toxins of Prymnesium parvum using a microalgal bioassay

Prymnesium parvum produces a variety of toxic compounds, which affect other algae, grazers and organisms at higher trophic levels. Here we provide the method for development of a sensitive algal bioassay using a microalgal target, Teleaulax acuta, to measure strain variability in P. parvum toxicity, as well as the temporal stability of both the intracellular and the extracellular lytic compounds of P. parvum. We show high strain variation in toxicities after 3 h incubation with LC₅₀s ranging from 24 to 223 × 10³ cells ml⁻¹. Most importantly we prove the necessity of testing physico-chemical properties of P. parvum toxins before attempting to isolate and characterize them. The extracellular toxin in the supernatant is highly unstable, and it loses significant lytic effects after 3 days despite storage at −20 °C and after only 24 h stored at 4 °C. However, when stored at −80 °C, lytic activity is more easily maintained. Reducing oxidation by storing the supernatant with no headspace in the vials significantly slowed loss of activity when stored at 4 °C. We show that the lytic activity of the intracellular toxins, when released by sonication, is not as high as the extracellular toxins, however the stability of the intracellular toxins when kept as a cell pellet at −20 °C is excellent, which proves this is a sufficient storage method for less than 3 months. Our results provide an ecologically appropriate algal bioassay to quantify lytic activity of P. parvum toxins and we have advanced our knowledge of how to handle and store the toxins from P. parvum so as to maintain biologically relevant toxicity.     

Continuous degradation of maltose by enzyme entrapment technology using calcium alginate beads as a matrix

Maltase from Bacillus licheniformis KIBGE-IB4 was immobilized within calcium alginate beads using entrapment technique. Immobilized maltase showed maximum immobilization yield with 4% sodium alginate and 0.2 M calcium chloride within 90.0 min of curing time. Entrapment increases the enzyme–substrate reaction time and temperature from 5.0 to 10.0 min and 45 °C to 50 °C, respectively as compared to its free counterpart. However, pH optima remained same for maltose hydrolysis. Diffusional limitation of substrate (maltose) caused a declined in V_max of immobilized enzyme from 8411.0 to 4919.0 U ml^?1 min^?1 whereas, K_m apparently increased from 1.71 to 3.17 mM ml^?1. Immobilization also increased the stability of free maltase against a broad temperature range and enzyme retained 45% and 32% activity at 55 °C and 60 °C, respectively after 90.0 min. Immobilized enzyme also exhibited recycling efficiency more than six cycles and retained 17% of its initial activity even after 6th cycles. Immobilized enzyme showed relatively better storage stability at 4 °C and 30 °C after 60.0 days as compared to free enzyme.     

Expression and purification of recombinant feline interferon in the baculovirus-insect larvae system

Feline interferons (FeIFNs) are cytokines with antiviral, antitumor and immunomodulatory functions used as therapeutic agents in a variety of veterinary diseases. In this work, FeIFN-α7 and FeIFN-α7xArg containing eight residues of arginine were expressed in Sf9 cells and insect larvae. At 4 days post-infection (dpi), the concentrations of FeIFN-α7 and FeIFN-α7xArg in suspension culture were (1.28 ± 0.15) × 10⁶ U ml⁻¹ and (1.3 ± 0.2) × 10⁶ U ml⁻¹ respectively. The maximum expression levels of FeIFN-α7 and FeIFN-α7xArg were (3.7 ± 0.2) × 10⁶ U ml⁻¹ and (3.5 ± 0.4) × 10⁶ U ml⁻¹ at 2 dpi in Rachiplusia nu larvae and (1.1 ± 0.2) × 10⁶ U ml⁻¹ and (1.0 ± 0.15) × 10⁶ U ml⁻¹ at 5 dpi in Spodoptera frugiperda larvae respectively. R. nu was a better host for FeIFN-α7 and FeIFN-α7xArg expression. The 8xArg tag did not affect the biological activity of FeIFN-α7 and was useful to promote the FeIFN-α7xArg adsorption on ion exchange chromatography (IEC), allowing its purification in a single step from supernatant culture and R. nu larvae. FeIFN-α7xArg was purified from the larval extract with a yield of 70% and a purification factor of 25 free of viruses. We conclude that R. nu larvae are new low-cost hosts for the expression of recombinant FeIFN-α7.     

Lethal effects of ichthyotoxic raphidophytes,Chattonella marina,C. antiqua,and Heterosigma akashiwo,on post-embryonic stages of the Japanese pearl oyster,Pinctada fucata martensii

The inimical effects of the ichthyotoxic harmful algal bloom (HAB)-forming raphidophytes Heterosigma akashiwo, Chattonella marina, and Chattonella antiqua on the early-life stages of the Japanese pearl oyster Pinctada fucata martensii were studied. Fertilized eggs and developing embryos were not affected following exposure to the harmful raphidophytes; however, all three algal species severely affected trochophores and D-larvae, early-stage D-larvae, and late-stage pre-settling larvae. Exposure to C. marina (5 × 10² cells ml⁻¹), C. antiqua (10³ cells ml⁻¹), and H. akashiwo (5 × 10³ cells ml⁻¹) resulted in decreased success of metamorphosis to the trochophore stage. A complete inhibition of trochophore metamorphosis was observed following exposure to C. antiqua at 5 × 10³ cells ml⁻¹ and C. marina at 8 × 10³ cells ml⁻¹. In all experiments, more than 80% of newly formed trochophores were anomalous, and in the case of exposure to H. akashiwo at 10⁵ cells ml⁻¹ more than 70% of D-larvae were anomalous. The activity rates of D-larvae (1-day-old) were significantly reduced following exposure to C. antiqua (8 × 10³ cells ml⁻¹, 24 h), C. marina (8 × 10³ cells ml⁻¹, 24 h), and H. akashiwo (10⁴ cells ml⁻¹, 24 h). The activity rates of pre-settling larvae (21-day-old) were also significantly reduced following exposure to C. antiqua (10³ cells ml⁻¹, 24 h), C. marina (8 × 10³ cells ml⁻¹, 24 h), and H. akashiwo (5 × 10⁴ cells ml⁻¹, 24 h). Significant mortalities of both larval stages were induced by all three raphidophytes, with higher mortality rates registered for pre-settling larvae than D-larvae, especially following exposure to C. marina (5 × 10²–8 × 10³ cells ml⁻¹, 48–86 h) and C. antiqua (10³–8 × 10³ cells ml⁻¹, 72–86 h). Contact between raphidophyte cells and newly metamorphosed trochophores and D-larvae, 1-day-old D-larvae, and 21-day-old larvae resulted in microscopic changes in the raphidophytes, and then, in the motile early-life stages of pearl oysters. Upon contact and physical disturbance of their cells by larval cilia, H. akashiwo, C. marina and C. antiqua became immotile and shed their glycocalyx. The trochophores and larvae were observed trapped in a conglomerate of glycocalyx and mucus, most probably a mixture of larval mucous and raphidophyte tricosyts and mucocytes. All motile stages of pearl oyster larvae showed a typical escape behavior translating into increased swimming in an effort to release themselves from the sticky mucous traps. The larvae subsequently became exhausted, entrapped in more heavy mucous, lost their larval cilia, sank, become immotile, and died. Although other toxic mediators could have been involved, the results of the present study indicate that all three raphidophytes were harmful only for motile stages of pearl oysters, and that the physical disturbance of their cells upon contact with the ciliary structures of pearl oyster larvae initiated the harmful mechanism. The present study is the first report of lethal effects of harmful Chattonella spp. towards larvae of a bivalve mollusc. Blooms of H. akashiwo, C. antiqua and C. marina occur in all major cultivation areas of P. fucata martensii during the developmental period of their larvae. Therefore, exposure of the motile early-life stages of Japanese pearl oysters could adversely affect their population recruitment. In addition, the present study shows that further research with early-life development of pearl oysters and other bivalves could contribute to improving the understanding of the controversial harmful mechanisms of raphidophytes in marine organisms.     

A novel low-temperature-active exo-inulinase identified based on Molecular-Activity strategy from Sphingobacterium sp. GN25 isolated from feces of Grus nigricollis

A novel glycosyl hydrolase family 32 exo-inulinase (InuAGN25) gene was cloned from Sphingobacterium sp. GN25 isolated from feces of Grus nigricollis. InuAGN25 showed the highest identity of 54.3% with a putative levanase recorded in GenBank. Molecular-Activity strategy was proposed to predict InuAGN25 to be a low-temperature-active exo-inulinase before experiments performance. Molecular analyses included progressive sequential, phylogenetic and structural analyses. InuAGN25 was effectively expressed in Escherichia coli. The purified recombinant InuAGN25 showed characteristics of low-temperature-active enzymes: (1) the enzyme retained 55.8% of the maximum activity at 20 °C, 35.8% at 10 °C, and even 8.2% at 0 °C; (2) the enzyme exhibited 75.8, 30.5 and 10.8% of the initial activity after preincubation for 60 min at 45, 50 and 55 °C, respectively; (3) K_m values of the enzyme toward inulin were 2.8, 3.0, 3.2 and 5.8 mg ml⁻¹ at 0, 10, 20 and 40 °C, respectively. Fructose was the main product of inulin and Jerusalem artichoke tubers hydrolyzed by the purified recombinant InuAGN25 at room temperature, 10 °C and 0 °C. These results suggested the Molecular-Activity strategy worked efficiently and made InuAGN25 promising for the production of fructose at low temperatures.     

Production of the postharvest biocontrol agent Bacillus subtilis CPA-8 using low cost commercial products and by-products

The aim of this research was to identify a low cost medium based on commercial products and by-products that provided maximum Bacillus subtilis CPA-8 growth and maintained biocontrol efficacy. Low cost media combining economical nitrogen and carbon sources such as yeast extract, peptone, soy products, sucrose, maltose and molasses were tested. Tests were carried out in 250-ml flasks containing 50 ml of each tested medium. Maximum cell growth (>3 × 10⁹ CFU ml^?1) was obtained in defatted soy flour 44% combined with sucrose or molasses media. Second, CPA-8 production was scaled up in a 5-l fermenter and CPA-8 population dynamics, pH and oxygen consumption in the optimized medium (defatted soy flour 44% – molasses) was recorded. In these tests, there was a 5-h lag phase before growth, after which exponential growth occurred and maximum production was 3 × 10⁹ CFU ml^?1 after 20 h. Fruit trials with cells and cell free supernatants from CPA-8 grown in optimized medium maintained biocontrol efficacy against Monilinia fructicola on peaches, resulting in disease reductions up to 95%. CPA-8 populations survived in wounds on inoculated peaches, regardless of the culture media used. The results show that B. subtilis CPA-8 can be produced in a low cost medium combining inexpensive nitrogen and carbon sources (40 g l^?1 defatted soy flour 44%, 5 g l^?1 molasses plus mineral trace supplements) in shake flasks and a laboratory fermenter (5 l). The results could be used to provide a reliable basis for scaling up the fermentation process to an industrial level.     

Management of white mold in processing tomatoes by Trichoderma spp. and chemical fungicides applied by drip irrigation

Field trials were carried out to evaluate six treatments combining biological agents and chemical fungicides applied via chemigation against white mold (Sclerotinia sclerotiorum) on processing tomatoes. The experiment was performed in Goiânia, Brazil, with tomato hybrid Heinz 7155 in 2009 and 2010 in a field previously infested with S. sclerotiorum sclerotia. Treatments were arranged in a randomized complete block design in a 2 × 3 factorial structure (with and without Trichoderma spp. 1.0 × 10⁹ viable conidia mL⁻¹ ha⁻¹) × fluazinam (1.0 L ha⁻¹), procimidone (1.5 L ha⁻¹) and control, applied by drip irrigation. Treatments were applied three times 10 days apart, starting one month after transplanting. Each treatment consisted of plots with three 72-meter rows with four plants m⁻¹ and 1.5 m spacing between rows, with three replications. Based on disease incidence evaluated weekly, the area under the disease progress curve (AUDPC) was obtained. Yield and its components were evaluated in addition to fruit pH and °Brix. Results were subjected to ANOVA, Scott-Knott (5%), and regression analysis. Biocontrol using Trichoderma spp. via chemigation singly or in combination with synthetic fungicides fluazinam and procimidone reduced AUDPC and increased fruit yield up to 25 t ha⁻¹. The best treatment for controlling white mold also increased pulp yield around 1.0 and 7.0 t ha⁻¹ in 2009 and 2010, respectively. The present work demonstrated the advantages of white mold biological control in processing tomato crops, where drip irrigation favored Trichoderma spp. delivery close to the plants and to the inoculum source.     

Biochemical characterization of a lichenase from Penicillium occitanis Pol6 and its potential application in the brewing industry

The purification and characterization of an extracellular lichenase from the fungus Penicillium occitanis Pol6 were studied. The strain produced the maximum level of extracellular lichenase (45 ± 5 U ml⁻¹) when grown in a medium containing oat flour (2%, w/v) at 30 °C for 7 days. The purified enzyme EG_L showed as a single protein band on SDS–PAGE with a molecular mass of 20 kDa. Its N-terminal sequence of 10 amino acid residues was determined as LDNGAPLLNV. The purified enzyme showed an optimum activity at pH 3.0 and 50–60 °C. The half-lives of EG_L at 60 °C and 70 °C were 80 min and 21 min, respectively. Substrate specificity studies revealed that the enzyme is a true β-1,3-1,4-d-glucanase. The enzyme hydrolyzed lichenan to yield trisaccharide, and tetrasaccharide as the main products. Under simulated mashing conditions, addition of EG_L (20 U/ml) or a commercial β-glucanase (20 U/ml) reduced the filtration time (25% and 21.3%, respectively) and viscosity (10% and 8.18%, respectively). These characteristics indicate that EG_L is a good candidate in the malting and brewing industry.     

Temperature-driven models of Aculops pelekassi (Acari: Eriophyidae) based on its development and fecundity on detached citrus leaves in the laboratory

This study was carried out to develop temperature-driven models for immature development and oviposition of the pink citrus rust mite Aculops pelekassi (Keifer). A. pelekassi egg development times decreased as the temperature increased, ranging from 6.6 days at 16 °C to 1.9 days at 35 °C. Total nymph development times decreased from 8.2 days at 16 °C to 3.3 days at 35 °C. The egg-to-adult development durations were 14.8, 11.6, 9.7, 8.0, 7.3, 6.1, and 5.2 days at 16, 20, 24, 26, 28, 32, and 35 °C, respectively. The lower developmental threshold temperatures for eggs, nymphs, and total egg-to-adult development were calculated as 9.3, 4.3, and 6.9 °C, respectively. The thermal constants were 54.0, 101.8, and 153.8 degree days for each of the above stages. The non-linear biophysical model fitted well for the relationship between the development rate and temperature for all stages. The Weibull function provided a good fit for the distribution of development times of each stage. Temperature affected the longevity and fecundity of A. pelekassi. Adult longevity decreased as the temperature increased and ranged from 24.2 days at 16 °C to 14.6 days at 35.0 °C. A. pelekassi had a maximum fecundity of 33.1 eggs per female at 28 °C, which declined to 18.8 eggs per female at 16 °C. In addition, three temperature-dependent components for an oviposition model of A. pelekassi were developed with sub-models estimated: total fecundity, age-specific cumulative oviposition rate, and age-specific survival rate. The oviposition model, coupled with the stage emergence model, should be useful to construct a population model for A. pelekassi in the future.     

Estudios de la viabilidad y la vitalidad frente al congelado de la levadura probiótica Saccharomyces boulardii: efecto del preacondicionamiento fisiológico

The aim of this study was to evaluate the vitality and viability of the probiotic yeast Saccharomyces boulardii after freezing/thawing and the physiological preconditioning effect on these properties. The results indicate that the specific growth rate (0.3/h^?1) and biomass (2-3 × 10⁸ cells/ml) of S. boulardii obtained in flasks shaken at 28 °C and at 37 °C were similar. Batch cultures of the yeast in bioreactors using glucose or sugar-cane molasses as carbon sources, reached yields of 0.28 g biomass/g sugar consumed, after 10 h incubation at 28 °C; the same results were obtained in fed batch fermentations. On the other hand, in batch cultures, the vitality of cells recovered during the exponential growth phase was greater than the vitality of cells from the stationary phase of growth. Vitality of cells from fed-batch fermentations was similar to that of stationary growing cells from batch fermentations. Survival to freezing at –20 °C and subsequent thawing of cells from batch cultures was 0.31% for cells in exponential phase of growth and 11.5% for cells in stationary phase. Pre-treatment of this yeast in media with water activity (a_w) 0.98 increased the survival to freezing of S. boulardii cells stored at –20 °C for 2 months by 10 fold. Exposure of the yeast to media of reduced a_w and/or freezing/thawing process negatively affected cell vitality. It was concluded that stress conditions studied herein decrease vitality of S. boulardii. Besides, the yeast strain studied presented good tolerance to bile salts even at low pH values.     

Thermal tolerance and preferred temperature range of juvenile meagre acclimated to four temperatures

The present study reports the temperature tolerance, estimated using dynamic and static methodologies, and preferred temperature range, based on oxygen consumption rate (OCR), of juvenile meagre (Argyrosomus regius) (Asso, 1801) (3.4±0.9 g) after 30 days of acclimation at 18, 22, 26 and 30 °C. Meagre has dynamic and static thermal tolerance zones of 551 °C² and 460 °C², respectively and is a low resistance fish species, with a resistance zone area of 87 °C². The OCR of juvenile meagre at the above acclimation temperatures was 370, 410, 618 and 642 mg h⁻¹ kg⁻¹, respectively, and is significantly different (P<0.0001, n=20). The fact that OCR increases by rising temperatures and gradually decreases after 26 °C indicates that the preferred temperature range of juvenile meagre is between 26 and 30 °C. Our study suggests that meagre is unable to respond to low and high temperature variation in aquaculture facilities or its natural habitats.     

Thermoregulation on the air–water interface—II: Foot conductance,activity metabolism and a two-dimensional heat transfer model

We used a quasi-adiabatic calorimeter and respirometry apparatus to measure heat loss from the feet of 3- to 4-d-old mallard ducklings (Anas platyrhynchos). We found that, at cool (<20 °C) operative temperatures, foot conductance increased in proportion to operative temperature, T_e, rather than water temperature. We combined these results with those of an earlier study to develop a heat transfer model for swimming ducklings. This model includes separate thermal conductances to air (0.027 W/°C-animal), to water through the down (0.035[1+2.05×10⁻⁷T_e⁴]) W/°C-animal, and to water through the feet (2.01×10⁻⁸T_e⁴ W/°C-animal). The overall conductance by all three routes is only 21% greater when swimming compared to standing in air at the same operative temperature. Interestingly, ducklings can maintain body temperature >39 °C while swimming in 5 °C water, but not when restrained in a calorimeter with 5 °C water. Peak oxygen consumption is greater when swimming, and apparently exercise metabolism substitutes almost completely for thermoregulatory heat production.     

Parameter estimation for a temperature-dependent development model of Thrips palmi Karny (Thysanoptera: Thripidae)

Development of immature Thrips palmi Karny was investigated at 12.5, 15, 17.5, 20, 22.5, 25, 27.5, 30, 32.5, and 35 °C, 20–40% RH and a photoperiod of 14:10 (L:D) h. Developmental time decreased with increasing temperature up to 32.5 °C in all stages. The total developmental time was longest at 12.5 °C (64.2 days) and shortest at 32.5 °C (9.2 days). The lower developmental threshold was 10.6, 10.6, 9.1, and 10.7 °C for egg, larva, prepupa, and pupa, respectively. The thermal constant required to complete the respective stage was 71.7, 59.2, 18.1, and 36.8DD. The lower threshold temperature and thermal constant were 10.6 °C and 183.3DD, respectively, for total immature development. The nonlinear relationship between developmental rate and temperature was well described by the modified Sharpe and DeMichele biophysical model (r² = 0.905–0.998). The distribution of developmental completion of each stage was described by the 3-parameter Weibull function (r² = 0.855–0.927). The temperature-dependent developmental models of T. palmi developed in this study could be used to predict its seasonal phenology in field and greenhouse vegetable crops.     

Direct in vivo interaction of the antibiotic primycin with the plasma membrane of Candida albicans: An EPR study

The direct interaction of the antibiotic primycin with the plasma membrane was investigated by employing the well-characterized ergosterol-producing, amphotericin B-sensitive parental Candida albicans strain 33erg⁺ and its ergosterol-less amphotericin B-resistant plasma membrane mutant erg-2. The growth inhibition concentration in shaken liquid medium was 64 μg ml^?1 for 33erg⁺ and 128 μg ml^?1 for erg-2, suggesting that the plasma membrane composition influences the mode of action of primycin. To determine the primycin-induced changes in the plasma membrane dynamic, electron paramagnetic resonance (EPR) spectroscopy methods were used, the spin-labeled fatty acid 5-(4,4-dimethyloxazolidine-N-oxyl)stearic acid) being applied for the in vivo measurements. The phase transition temperatures of untreated strain 33erg⁺ and its mutant erg-2 were 12.5 °C and 11 °C, respectively. After 128 μg ml^?1 primycin treatment, these values increased to 17.5 °C and 16 °C, revealing a significant reduction in the phospholipid flexibility. Saturation transfer EPR measurements demonstrated that, the rotational correlation times of the spin label molecule for the control samples of 33erg⁺ and erg-2 were 60 ns and 100 ns. These correlation times gradually decreased on the addition of increasing primycin concentrations, reaching 8 μs and 1 μs. The results indicate the plasma membrane “rigidizing” effect of primycin, a feature that may stem from its ability to undergo complex formation with membrane constituent fatty acid molecules, causing alterations in the structures of phospholipids in the hydrophobic surface near the fatty acid chain region.     

Emulsifying properties of a marine bacterial exopolysaccharide

Exopolysaccharide produced by a marine Enterobacter cloaceae (designated as EPS 71a) emulsified hexane, benzene, xylene, kerosene, paraffin oil, cottonseed oil, coconut oil, jojoba oil, castor oil, groundnut oil and sunflower oil. However, it could form stable emulsions with groundnut oil and hexane at optimal concentration of 1 mg ml⁻¹. Further increase in concentration of EPS 71a did not show noteworthy increase in emulsification indices. Emulsions with groundnut oil and hexane were stable up to 10 days between pH 2 and 10 and in the presence of sodium chloride in the range of 5–50 mg ml⁻¹ at 35–37 °C. EPS 71a formed stable emulsion with xylene as compared to commercial gums such as arabic, tragacanth, karaya and xanthan.     

Spore production in solid-state fermentation of rice by Clonostachys rosea,a biopesticide for gray mold of strawberries

Gray mold caused by Botrytis cinerea is an important disease of strawberry. Clonostachys rosea is a mycoparasite of B. cinerea that reduces fruit losses when used as a biocontrol agent. Since spore production by C. rosea has not been optimized, we investigated factors affecting sporulation under aseptic conditions on white rice grains. The greatest spore production in glass flasks, 3.4 × 10⁹ spores/g-dry-matter (gDM), occurred with an initial moisture content of 46% (w/w wet basis), inoculated with 1 × 10⁶ spores/gDM and hand shaken every 15 days. However, a lower inoculum density (9 × 10³ spores/gDM) and no shaking also gave acceptable sporulation. In plastic bags 1.1 × 10⁸ spores/gDM were produced in 15 days, suggesting that larger scale production may be feasible: with this spore content, 24 m² of incubator space would produce sufficient spores for the continued treatment of 1 ha of strawberry plants.