Phacidiopycnis washingtonensis,Cause of a New Storage Rot of Apples in Northern Europe

Phacidiopycnis washingtonensis was identified by morphology and ITS sequence analysis as the cause of rubbery rot, a new storage disease of apples in northern Germany. Infected fruits had an unusually firm texture and pale appearance after storage in ultra‐low oxygen conditions, but turned dark brown to black in ambient atmosphere. Ultimately, the surface of rotted fruits became covered by black pycnidia producing cream‐coloured conidial exudates. Rubbery rot affected several apple varieties, including the commercially important ‘Jonagold’ and ‘Elstar’. Losses during storage were commonly below 1% but reached 5–10% in a few cases. Fruits of ‘Golden Hornet’ crab apple trees planted as pollinators in commercial orchards became heavily infected by P. washingtonensis in October. Conidia were released throughout the following season from infected fruit mummies, which remained attached to the crab apple tree.     

Virulence and the Production of Endo-1,4-β-glucanase by Isolates of Alternaria alternata Involved in the Moldy-core Disease of Apples

Alternaria alternata, is the predominant fungal pathogen responsible for moldy‐core in apple cultivars of the Red Delicious group. Here we report on the association between virulence of natural isolates of A. alternata, their production of endo‐1,4‐β‐glucanase (EG) and moldy‐core development in apple fruits. Based on decay development following wound inoculations of mature fruits, three of 150 isolates, collected in three orchards in northern Israel and representing low, moderate and high virulence, were selected and designated Rm44, Er30 and Sh42, respectively. All three isolates secreted EG when grown on enzyme‐inducing medium (EIM) containing commercial cellulose or apple cell walls and this production was related to their degree of virulence. Polyacrylamide gel electrophoresis (PAGE) revealed quantitative differences between the three isolates, relative to their virulence. When fungal extracts were run in native gels, a single band with a molecular mass of 23 kDa showing EG activity was produced by the high‐ (Sh42) and the medium‐virulence (Er30) isolate but not by the low‐virulence (Rm44) isolate. A commercial cellulase preparation (containing endo‐ and exo‐1,4‐β‐glucanase) placed on pricked fruit led to the formation of symptoms similar to those developing on A. alternata‐inoculated fruits within 2–4 days. Inoculation of bloom clusters at full bloom with the highly virulent isolate (Sh42) of A. alternata resulted in a significantly higher infection in fruits (58%) than in those inoculated with the low‐virulence isolate (Rm44) (30%). Our results suggest that the moldy‐core symptoms caused by A. alternata in apple, could be related to the ability of the fungus to produce EG in developing lesions.     

Effect of Cultivation Time on Production of Antifungal Metabolite(s) by Streptomyces hygroscopicus in Laboratory‐Scale Bioreactor

Biological control agents offer one of the best alternatives to reduce the use of pesticides . Fungi from the genera Alternaria, Colletotrichum and Fusarium are listed among the most important storage pathogens of apple fruits. During storage, transport and marketing, pathogenic fungi can cause significant losses of apple fruits. This investigation studied the potential of Streptomyces hygroscopicus as a biocontrol agent against pathogenic fungi obtained from apple fruit samples expressing rot symptoms. Production of antifungal metabolites by S. hygroscopicus was carried out in 3‐l bench‐scale bioreactor (Biostat^® Aplus, Sartorius AG, Germany) during 7 days. Fermentation was carried out at 27°C with aeration rate of 0.5 vvm and agitation rate of 200 rpm. The aim was to analyse bioprocess parameters of batch biofungicide production in medium containing glucose as a carbon source and to examine at which stage of bioprocess production of antifungal metabolite(s) against six phytopathogenic fungi occurs. In vitro antifungal activity of the produced metabolites against six fungi of the genera Colletotrichum, Fusarium and Alternaria grown on potato dextrose agar were determined every 24 h using wells technique. Antifungal activity of cell‐free culture filtrate and filtrate treated with high temperature were tested. The filtrate treated with high temperature did not show any antifungal activity suggesting that active components are thermo unstable. Stationary phase of growth occurred between the third and fourth day of cultivation when production of secondary metabolites begins. Obtained results showed that maximal antifungal activity is achieved on fifth and sixth day of S. hygroscopicus cultivation under defined conditions (inhibition zone diameter higher than 30 mm for all test fungi).     

Patulin and aflatoxin in brown rot lesion of apple fruits and their regulation

Aspergillus flavus, A. niger, Penicillium expansum and Rhizopus stolonifer were the most frequently isolated fungi from healthy apple fruits. Alternaria alternata was the most common organism of rotten apple fruits, followed by A. niger, A. flavus, P. expansum and R. stolonifer. The prevalent type of decay, brown rot lesion, is caused by R. stolonifer followed by A. flavus, A. niger, A. alternata and P. expansum. Sodium hypochlorite had good curative properties against fruit rots. The main natural mycotoxins produced in rotten apple were patulin and aflatoxins. The optimum temperature for patulin production by P. expansum was 15 °C after 15 days. Complete inhibition of patulin formation was attained using 0.2% lemon oil and > 90% inhibition using 0.05% lemon and 0.2% orange oils. Also significant inhibition (> 90%) of aflatoxin production was observed with 0.2% lemon oil. This revised version was published online in July 2006 with corrections to the Cover Date.     

Fusarium Head Blight of Common Polish Winter Wheat Cultivars – Comparison of Effects of Fusarium avenaceum and Fusarium culmorum on Yield Components

The effects of Fusarium avenaceum and Fusarium culmorum on the reduction in yield components, after independent inoculation of 14 winter wheat cultivars, were investigated. Single isolates of F. avenaceum and F. culmorum were independently used in inoculations of winter wheat heads. Reductions in the following yield traits: 1000‐kernel weight (TKW), the weight (WKH) and number (NKH) of kernels per head after inoculation were analysed statistically. The results indicate differences between both pathogens in their effects on yield traits. The statistical calculations were performed using analysis of variance (a three‐factor experiment) for particular yield trait reductions and multivariate analysis of variance for the yield trait reductions jointly. Almost all of the univariate and multivariate hypotheses concerning no differences between pathogens (F. culmorum, F. avenaceum), climatic conditions (years) and cultivars as well as hypotheses concerning no interactions between factors (pathogens, years, cultivars) were rejected at least at P= 0.05 significance level. The reduction of yield traits indicated individual reactions of the tested winter wheat cultivars to different pathogens. Among the tested traits the highest influence on the rejection of the hypothesis concerning the equivalence of F. avenaceum and F. culmorum was observed for TKW and WKH. The effect of the pathogen on yield reduction was greater for F. avenaceum than for F. culmorum during 1996 and 1997. A comparison of the cultivars indicated that the Begra cultivar showed the highest tolerance to inoculation with both Fusarium pathogens. Moreover, this genotype as well as several others showed lower tolerance to F. avenaceum rather than to F. culmorum, whereas Elena was the only cultivar with the opposite tendency.     

Lower Temperatures More Effective than Atmosphere Modification in Controlling Botrytis and Nectria Rots in Stored Apples

A series of small‐scale controlled inoculation experiments has been conducted during 2005–2009 to determine whether temperature and controlled atmosphere (CA) storage conditions affect significantly the incidence of Botrytis cinerea and Neonectria galligena rots of apples and to assess whether CA regimes can be ‘fine‐tuned’ to suppress fungal rotting. The incidence of B. cinerea and N. galligena rots on apple was reduced consistently by storage in lower temperatures (1.5–2°C). In no case was the disease incidence significantly higher than that under air storage conditions. However, the effect of CA conditions on rot development varied greatly from year to year so that overall there were no significant effects of CA conditions on the incidence of rot during storage till the following April. Thus, we can neither fine‐tune CA conditions to reduce rot development nor exploit the advantages of lower storage temperatures for Cox and Bramley for this purpose due to inevitable development of low temperature breakdown. Further research is needed to study the effects of ethylene control technologies and modulated storage temperatures on rot development.     

Mycotoxins produced by toxic Fusarium isolates obtained from agricultural and nonagricultural areas (Arctic) of Norway

Twenty-five isolates of F. acuminatum, 38 of F. avenaceum, 1 of F. culmorum, 31 of F. oxysporum and 56 of F. sambucinum were obtained in 1983, 1984 and 1986 from cereal grains and soil from various parts of Norway. The isolates were grown on an autoclaved Uncle Ben's parboiled rice medium and examined for production of trichothecenes and other toxins and for toxicity in rat feeding tests. F. culmorum N46C(2) and Fusarium sambucinum 45-86-A produced zearalenone (F-2) 864 and 665 ppm, respectively and caused uterine enlargement in rats. Most of these isolates produced no known trichothecene mycotoxins that could account for the toxicity that was demonstrated in the rat feeding tests. All but F. avenaceum N26B produced fusarin C (1.5 ppm) but caused no toxic effects in rat feeding test. None of the isolates produced fusarochromanone (TDP-1). Thirteen isolates of F. acuminatum, 16 of F. avenaceum, 14 of F. oxysporum and 3 of F. sambucinum produced a cytotoxic factor which we named HM-8. One isolate of F. avenaceum, 12 of F. oxysporum and 46 of F. sambucinum produced a hemorrhagic factor which we named H-1 (wortmannin). Twenty isolates of F. acuminatum, 22 of F. avenaceum, 17 of F. oxysporum and 1 of F. sambucinum produced moniliformin. Four isolates of F. acuminatum, 9 of F. avenaceum, 25 of F. oxysporum and 52 of F. sambucinum caused death to rats. Three isolates of F. avenaceum, 19 of F. oxysporum and 47 of F. sambucinum induced hemorrhage in various organs. All isolates caused decreased weight gain, relative to the control diets.     

Occurrence and Latent Infection of Alternaria Rot of Pingguoli Pear (Pyrus bretschneideri Rehd. cv. Pingguoli) Fruits in Gansu, China

Alternaria rot of Pingguoli pear occurred after latent infection. Fruit surfaces were asymptomatic within 60 days storage under cold condition (0°C, RH 85–90%), but black‐grey hyphae could be seen in the lenticels or calyx tube of Pingguoli pear after 90 days of storage. The tissue collapsed and resulted in visible black spots as the hyphae spread over the fruit. Average incidence of Alternaria rot of fruits from an orchard in Gansu was 28.86% at 100 days of storage. The main fungus isolated from the Alternaria rot on stored Pingguoli pear was identified as Alternaria alternata (Fr. : Fr.) Keissl. This pathogen was able to initially infect the fruit via two pathways during the growing season, and then remain in a latent state. The fungus first colonized the styles at the full‐blossoming stage, and then grew into the carpel cavities progressively after 50 days from petal fall. The percentage latent infection of A. alternata was up to 45% in the carpel cavity until the harvest time. The fungus also attacked fruit surfaces and remained latent in the fruit peel during fruit development. The percentage of A. alternata latent infection at the calyx end, middle part and stem end of the fruit peel was 40%, 24% and 42.8%, respectively, at harvest time.     

Identification of Colletotrichum Species Causing Bitter Rot of Apple and Pear in Croatia

Symptoms of bitter rot were observed on apple and pear fruits in the field and in storage in Croatia between 2009 and 2011. Fifteen Colletotrichum isolates from apple and two from pear were collected and identified by sequencing of the ITS1 and ITS2 regions of the ribosomal DNA. Phylogenetic analysis revealed that ten isolates from apple and two isolates from pear could be identified as Colletotrichum fioriniae, five isolates from apple were clustered in Colletotrichum clavatum, while one isolate was in the Colletotrichum acutatum A7 group. All isolates caused typical bitter rot symptoms when inoculated on apple and pear fruits.     

Effects of organomercury treatment of wheat seed on Fusarium seedling disease in inoculated soil

In growth room experiments, treatment of wheat seed with phenyl mercuric acetate significantly reduced seedling disease caused by soil-borne Fusarium culmorum, F. nivale and F. avenaceum in inoculated soil. Seed treatment improved the low germination caused by F. culmorum, but did not affect the reduced seedling vigour caused by F. nivale. Seed treatment was normally as effective in conditions favouring disease as it was in less favourable conditions. Conditions favouring disease in soil inoculated with F. culmorum were high inoculum level, deep sowing and dry soil.     

苹果渣乙醇提取物抗菌活性及防腐作用研究

为提高苹果渣资源利用率,探究苹果渣乙醇提取物的抗菌活性和防腐性能,该文采用微波辅助提取法制取苹果渣乙醇提取物,用抑菌圈实验测定其抗菌活性,并研究了其防腐作用。结果表明:(1)苹果渣乙醇提取物对酵母菌抑制作用不明显(抑菌圈直径<1 mm),对金黄色葡萄球菌和大肠杆菌的抑菌作用较明显(抑菌圈直径为6～9 mm),最佳抑菌浓度为4.0 g·L^-1。(2)pH值和盐浓度对其抑菌效果有影响,pH值为6～7,盐浓度为5.0 g·L^-1,抑菌效果最好。(3)对百香果有较好的保鲜防腐效果,最佳使用浓度为0.2%。在该浓度下贮藏后的百香果腐烂率为6.7%(对照组为67%),失重率为5.5%(对照组为36.3%),可溶性固形物、总酸含量均与贮藏前差异不显著(P> 0.05)(对照组P< 0.05),且果实较饱满,硬度较高,鲜艳有光泽,酸甜适中。综上所述,苹果渣乙醇提取物对大肠杆菌和金黄色葡萄球菌具有良好的抑制作用,对百香果的保鲜防腐效果佳,可应用到天然食品的保鲜防腐。     

First report of Alternaria leaf spot caused by Alternaria alternata (Fries.) Kiessler on Sonchus oleraceus L. and Convolvulus arvensis L. in Algeria

Alternaria leaf spot caused by Alternaria alternata (Fries.) Kiessler was found on sow thistle (Sonchus oleraceus L.) and field bindweed (Convolvulus arvensis L.) in the experimental station of ENSA (Ecole Nationale Supérieure d'Agronomie) in Algiers, Algeria, during the winter of 2016. Necrotic spots in the form of concentric circles were observed on the leaves of both weeds with disease incidence of approximately 70% and disease severity ranging from 50% to 70%. Fungi were isolated from the infected leaves and identified as A. alternata, based on morphological and molecular analyses (using genetic marker internal transcribed spacer, ITS of rDNA). Pathogenicity tests confirmed that A. alternata is the causing agent of leaf spot disease of sow thistle and field bindweed in accordance with the original symptoms. To the best of our knowledge, this is the first report of sow thistle and field bindweed naturally infected by A. alternata in Algeria.     

Control of apple blue mold by <Emphasis Type="Italic">Pichia pastoris</Emphasis> recombinant strains expressing cecropin A

Recombinant Pichia pastoris yeasts expressing cecropin A (GS115/CEC), was evaluated for the control of the blue mold of apple caused by Penicillium expansum due to cecropin A peptide’s effective antimicrobial effects on P. expansum spores by the thiazolyl blue (MTT) assay. Then, the protein concentration was determined and it was expressed at high levels up to 14.2 mg/L in the culture medium. Meanwhile, the population growth was assayed in vivo. The population growth of recombinant strain GS115/CEC was higher than that of non-transformed strain GS115 in red Fuji apples wounds. Recombinant yeast strains GS115/CEC significantly inhibited growth of germinated P. expansum spores in vitro and inhibited decay development caused by P. expansum in apple fruits in vivo when compared with apple fruits inoculated with sterile water or the yeast strain GS115/pPIC (plasmid pPIC9k transformed in GS115). This study demonstrated the potential of expression of the antifungal peptide in yeast for the control of postharvest blue mold infections on pome fruits.     

2-hydroxy-3-phenylpropanoic acid suppressed the growth of Alternaria alternata through damaging cell membrane integrity and modulating reactive oxygen species metabolism

Black spot rot caused by Alternaria alternata is one of the major postharvest disease of apple fruit during logistic. This study evaluated in vitro inhibitory effect of 2-hydroxy-3-phenylpropanoic acid (PLA) at various concentrations on A. alternata and the possible mechanisms involved in its action. Results showed that different concentrations of PLA inhibited conidia germination and mycelial growth of A. alternata in vitro, and 1.0 g L⁻¹ was the lowest effective concentration to suppress A. alternata growth. Moreover, PLA significantly reduced relative conductivity and increased malondialdehyde and soluble protein contents. PLA also increased H₂O₂ and dehydroascorbic acid contents, but reduced ascorbic acid content. Additionally, PLA treatment inhibited catalase, ascorbate peroxidase, monodehydroascorbate acid reductase, dehydroascorbic acid reductase and glutathione reductase activities, whereas promoted superoxide dismutase activity. All these findings suggest that the possible mechanisms involved in the inhibitory effect of PLA on A. alternata included damaging the cell membrane integrity to cause electrolyte leakage and destroying reactive oxygen species balance.     

Influence of pest control pressure on occurrence of ground beetles (Coleoptera: Carabidae) in apple orchards

Bioindicators that reflect the level of chemical control have not been developed for use in apple orchards in Japan. In Japan, many carabid beetles and spiders are beneficial predators of a wide range of agricultural pest insects. In apple orchards, spider numbers are reflective of the level of control pressure, but there is no information on carabid beetles. To establish whether carabid beetles could indeed be useful bioindicators, the number of carabid beetles and spiders in six apple orchards (A–F) under different levels of control pressure was investigated in Akita prefecture, northern Japan, in 2008 and 2009. The negative impact on beneficial arthropods was assumed to decrease in the order of pyrethroids (A) > organophosphates (B) > neonicotinoids (C) > insect growth regulators (D) > fungicides only (E) > no spraying (F). Sprays were applied at 2-week intervals from mid-May to early August. Twenty species of carabid beetles were found in orchard F; about 50% of adults were Anisodactylus punctatipennis, and 30–40% were Amara chalcites. The number of Am. chalcites significantly decreased with increasing control pressure: (A–C) < (E, F) in 2008, (A–E) < F in 2009, and (A, B) < E in 2009. Most spiders collected were juvenile Tetragnathidae and Araneidae. The numbers of spiders and Am. chalcites adults were significantly highly correlated at each control pressure in both years. Thus, it was considered that only Am. chalcites adults can be used as bioindicators reflecting the control pressure in apple orchards in northern Japan, whereas the main diet of this species is probably C3 plants.     

A method for pre-harvest assessment of latent infections in fruits

A method was developed for the assessment of latent infections of Alternaria alternata in mango fruits. Fruit discs were sampled and assayed for fungal contamination under suitable growth conditions. By this method it was possible to evaluate the percentage of infections and the relative surface infected by Alternaria in mango fruits before harvest. The percentage of infections increased during the growing season but fungicide treatments prevented this increase. A significant positive correlation was found between the relative surface infected by latent Alternaria at harvest time and the incidence of black spots that developed on the fruit during storage.     

Potential host fruits for Drosophila suzukii: olfactory and oviposition preferences and suitability for development

Drosophila suzukii (Matsumura) (Diptera: Drosophilidae), the spotted wing drosophila, is a pest endemic to Southeast Asia that invaded the Americas and Europe in 2008. In contrast to most of its congeners, D. suzukii possesses a serrated ovipositor that allows it to lay eggs in unwounded commercial fruits, resulting in severe revenue losses for the industry. The purpose of this study was to determine the susceptibility of known host fruits, including cherry, strawberry, blueberry, and grape, and potential host fruits, such as banana and apple, to attack by D. suzukii. Based on the responses to volatile cues offered in a six‐choice olfactometer, the preference of female D. suzukii was ranked in the following order: strawberry = cherry > banana = apple = blueberry = grape, but in no‐choice and choice oviposition tests, the preferences were ranked as follows: cherry > strawberry = blueberry > grape = banana > apple. Furthermore, we reconfirmed that D. suzukii mainly targets rotten fruit for feeding and ripe fruit for oviposition, and females preferred fruits with intensive mechanical damage. Based on developmental parameters, apple was the least suitable host. This study has implications for the control of D. suzukii, especially in mixed fruit orchards, by providing a promising avenue for exploiting behaviour‐based control tools and emphasizing the importance of phenology in host fruit susceptibility.     

Cryptolaemus montrouzieri as a predator of the striped mealybug,Ferrisia virgata,reared on two hosts

The striped mealybug, Ferrisia virgata (Cockerell) (Hemiptera: Pseudococcidae), is a cosmopolitan pest of a variety of agricultural crops including cotton. To investigate the biological control potential of the predatory ladybird Cryptolaemus montrouzieri Mulsant (Coleoptera: Coccinellidae) against this pest, we evaluated its developmental and reproductive fitness when feeding on F. virgata reared on pumpkin fruits or on cotton leaves and compared this to a diet of Planococcus citri Risso (Hemiptera: Pseudococcidae) reared on pumpkin fruits. F. virgata and P. citri reared on pumpkins were equally suitable prey for the pre‐imaginal stages of C. montrouzieri. Duration of total immature development was 1 day longer in C. montrouzieri offered F. virgata reared on cotton as compared with F. virgata or P. citri reared on pumpkin, whereas no significant difference was observed in survival rates. Diet significantly influenced the reproductive fitness of C. montrouzieri. Females offered P. citri reared on pumpkin had significantly shorter pre‐oviposition periods and higher fecundity and fertility than those given F. virgata reared on pumpkin or cotton leaves. F. virgata grown on cotton leaves supported the reproduction of C. montrouzieri better than F. virgata reared on pumpkin. Our study established that C. montrouzieri can successfully complete its development and reproduction when fed exclusively on F. virgata and indicates its potential as a biological control agent of this emerging cotton pest.     

Molecular characterization of Alternaria alternata population isolated from Upper Egyptian tomato fruits

Alternaria alternata is the most common fungal pathogen of tomatoes in Upper Egypt. Morphological identification of this fungus is challenging; therefore, this study searched for new classification tools based on molecular techniques. Using a dilution plating method, 67 strains of A. alternata were isolated from 34 samples of rotten tomato fruits representing the Giza 80 and Edkawy cultivars. The collected strains were identified using the amplification products of the internal transcribed spacer (ITS) region, glyceraldehyde 3‐phosphate dehydrogenase (Gpd) and Alt a1, which is a gene involved in the production of most of the allergens produced by A. alternata. The screening revealed that A. alternata constituted more than half of the total fungi recovered from rotten tomatoes in this study. According to the phylogenetic analysis using these three loci, the collected strains clustered in accordance with the host cultivar type from which they had been isolated. Specific gene random primer polymerase chain reaction (SGRP‐PCR) techniques indicated that the A. alternata population in the tested region has a high genetic diversity. The pathogenicity test showed that most of the A. alternata isolates (67.2%) were highly pathogenic, and no correlation was found between the phylogenetic analysis and pathogenicity. In addition, the influence of the fungicide Disan 80% on the collected strains showed significant differences that were attributed to the source of isolation.     

Role of Pear Fruit Cuticular Wax and Surface Hydrophobicity in Regulating the Prepenetration Phase of Alternaria alternata Infection

The role of cuticular wax and the surface hydrophobicity of the fruit of the ‘Zaosu’ pear (Pyrus bretschneideri Rehd) in regulating the prepenetration phase of Alternaria alternata infection were analysed in vivo and in vitro. Results showed that cuticular wax on an intact fruit surface, as well as wax extracts mounted on silanized glass slides or onion epidermis, favoured the formation of short, differentiated germ tubes and large numbers of appressoria (APP) or infected hyphae (IH). Dewaxed fruits or no wax extract mounted on in vitro surfaces, however, enhanced germ tube elongation and inhibited or delayed the formation of infection structure. High surface hydrophobicity resulting from cuticular wax also stimulated infection structure formation, as contact angle (hydrophobicity) was positively correlated with APP formation but negatively correlated with germ tube elongation. Alternaria alternata cutinase enzyme activity was also induced by cuticular wax, both in vivo and in vitro. These findings suggest that the chemical composition and hydrophobicity of pear fruit cuticular wax are essential in facilitating fungal invasion by regulating the growth and differentiation of A. alternata during the prepenetration phase.