Characterization of Neofabraea vagabunda isolates causing apple bull's eye rot in Italy (Emilia‐Romagna region)

Neofabraea vagabunda is the prevalent cause of bull's eye rot, one of the main postharvest diseases of apple, in many producing areas, but its biology has not been studied in detail. The molecular identification, by DNA sequencing of the β‐tubulin region, of 41 isolates collected from apples showing bull's eye rot in the Emilia‐Romagna region confirmed N. vagabunda as the main species in Italy. A biological and morphological characterization of N. vagabunda isolates was performed in vitro. Assays at temperatures ranging from 0 to 30 °C carried out on 10 isolates demonstrated: (i) a marked influence of temperature on colony morphology, conidial production, conidial size and mycelial growth, showing the cold‐tolerant character of N. vagabunda; and (ii) that culture at 15 °C on tomato agar (TA) for 14 days is a rapid and reliable method to favour pathogen conidial production. Trials performed on 38 isolates using these incubation conditions recorded the presence of two N. vagabunda morphotypes, differing for colony morphology, conidial size, conidiomata formation and temperature requirement. The alkalizing ability of the pathogen during growth on TA was also demonstrated for the first time. The pathogenicity of 25 N. vagabunda isolates was proved in vivo on artificially infected Cripps Pink apples. A pH increase was also recorded in apple tissue infected by N. vagabunda isolates (on average 0.2 and 0.3 units of pH after 60 and 120 days of incubation, respectively), suggesting that the N. vagabunda transition from quiescence to necrotrophic colonization in apples could involve the secretion of alkalizing compounds.     

Effect of hot water treatment on peach volatile emission and Monilinia fructicola development

The effect of hot water treatment (HWT) to control peach brown rot was investigated. Peaches were dipped in water at 60°C for 60 s and artificially inoculated with Monilinia fructicola conidia. HWT failed to control brown rot if applied before inoculation and microscopic observations revealed a stimulatory effect on germ tube elongation of M. fructicola conidia placed immediately after HWT on the fruit surface, compared to the control. The influence of fruit volatile emission due to HWT was performed on the pathogen conidia exposed to the headspace surrounding peaches. The results showed an increase of M. fructicola conidial germination ranging from 33 to 64% for cultivars Lucie Tardibelle and Red Haven heat‐treated peaches, respectively, compared to the control. The volatile blend emitted from heat‐treated fruit was analysed by solid‐phase microextraction/gas chromatography‐mass spectrometry (SPME/GC‐MS) and proton transfer reaction‐time of flight‐mass spectrometry (PTR‐ToF‐MS). Fifty compounds were detected by SPME/GC‐MS in volatile blends of cv. Lucie Tardibelle peaches and significant differences in volatile emission were observed among heated and control fruit. Using PTR‐ToF‐MS analysis, acetaldehyde and ethanol were detected at levels 15‐ and 28‐fold higher in heated fruit compared to unheated ones, respectively. In vitro assays confirmed the stimulatory effect (60 and 15%) of acetaldehyde (0·6 μL L^?1) and ethanol (0·2 μL L^?1) on M. fructicola conidial germination and mycelial growth, respectively. The results showed that volatile organic compounds (VOCs) emitted from heat‐treated peaches could stimulate M. fructicola conidial germination, increasing brown rot incidence in treated peaches when the inoculation occurs immediately after HWT.     

Effect of apple cultivars and storage periods on the virulence of Neofabraea spp.

Bull’s eye rot is a typical quiescent postharvest apple disease in major fruit-growing areas. The susceptibility of different apple cultivars to Neofabraea spp. (N. vagabunda and N. malicorticis) was assessed, with Granny Smith showing the most resistance and Cripps Pink the most susceptibility. To assess the factors involved in conidial germination, Neofabraea spp. were grown on crude protein extracts (CPEs) collected from apple fruits at different storage periods. Fungal germ tube growth rate and pathogenic enzyme (cellulase and xylanase) activity were assessed. Results showed that CPEs collected after 2 and 4 months of storage progressively stimulated conidial germination and germ tube elongation, while a lesser effect was observed from CPEs after 1 month of storage. Xylanase proved to be the main degrading enzyme secreted by all the isolates, while cellulase was produced only by N. vagabunda isolates. Overall, the isolate ID02 was the most virulent, based on more rapid germ tube elongation and greater activity of the lytic enzymes.     

Role of strawberry volatile organic compounds in the development of Botrytis cinerea infection

Botrytis cinerea, the main pathogen of strawberry, has the ability to remain quiescent in unripe tissue and develop disease symptoms in ripe fruit. As strawberry ripening is characterized by an increase of aroma compounds, the role of volatile emission in the development of infection was investigated. Thirty‐five strawberry volatile organic compounds (VOCs) were tested on B. cinerea in vitro and volatile emission was analysed in strawberry harvested at four ripening stages by headspace solid‐phase microextraction/gas chromatography–mass spectrometry and proton transfer reaction–time of flight–mass spectrometry. The coupling of such data sets made it possible to conclude that key strawberry aroma compounds stimulate B. cinerea conidial germination and some typical wound‐volatiles stimulate pathogen conidial germination or mycelial growth. This study is the first report of fungal stimulation by some VOCs naturally occurring in strawberry: the esters ethyl butanoate, cis‐3‐hexenyl acetate, trans‐2‐hexenyl acetate, methyl butanoate and hexyl butanoate, the furanones furaneol and mesifurane, and the alcohol trans‐2‐hexenol. The results of this work provide advances in understanding the functional role of fruit VOCs and suggest, for the first time, that fruit VOCs may influence the development of B. cinerea from the latent phase and that they could favour the invasive growth of B. cinerea after wounding. In particular, ethyl butanoate and furaneol could signal strawberry ripening, and the green leaf volatiles trans‐2‐hexenol, trans‐2‐hexenyl acetate and cis‐3‐hexenyl acetate could signal the presence of damaged tissues that are easier sites for penetration by B. cinerea.     

Phenotypic,molecular and pathogenic characterization of Phlyctema vagabunda,causal agent of olive leprosy

Olive leprosy, caused by the fungus Phlyctema vagabunda, is a classic fruit rot disease widespread in the Mediterranean basin. From 2009 to 2013, new disease symptoms consisting of small circular necrotic leaf lesions, coin branch canker and shoot dieback were observed in Spanish and Portuguese olive orchards showing intense defoliation. Phlyctema‐like anamorphs were consistently isolated from leaves and shoots with symptoms. Representative isolates from affected leaves, shoots and fruits were characterized based on morphology of colonies and conidia, optimum growth temperature and comparison of DNA sequence data from four regions: ITS, tub2, MIT and rpb2. In addition, pathogenicity tests were performed on apple and olive fruits, and on branches and leaves of olive trees. Maximum mycelial growth rate ranged between 0.54 and 0.73 mm per day. Conidia produced on inoculated apple fruits showed slight differences in morphology among the representative fungal isolates evaluated. Phylogenetic analysis clustered all of the Phlyctema‐like isolates in the same clade, identifying them as Phlyctema vagabunda. On fruits, influence of wounding, ripening and cultivar resistance was studied, with cv. Blanqueta being the most susceptible cultivar. On branches, a mycelial‐plug inoculation method reproduced olive leprosy symptoms and caused shoot dieback. On leaves, Koch's postulates were fulfilled and the pathogen caused characteristic necrotic spots and plant defoliation. This is the first time that the pathogenicity of P. vagabunda in olive leaves has been demonstrated.     

<Emphasis Type="Italic">Neofusicoccum parvum</Emphasis> associated with fruit rot and shoot blight of peaches in Greece

Shoot blights and fruit rots comprise the most serious diseases of peaches in Greece. In this study, the importance of the fungus Neofusicoccum parvum as a casual agent of a fruit rot and shoot blight of peach trees in Greece was investigated. This pathogen was isolated from both immature and mature peach fruit of the cultivar “Catherine” and later on from mature fruit of the peach cultivars “Andross”, “RedHaven”, “Sun Crest” and “Sun Cloud”. In the first year of investigation, N. parvum was found causing preharvest fruit rot and shoot blights of peach trees only at the location “Ammos-Mesi-Meliki Verias” in the prefecture of Imathia (the main peach production area of Greece) at incidences of 30 and 8%, respectively. However, in 2006 N. parvum was isolated from more locations such as Diavatos, Veria, Kopanos and Agia Marina in the prefecture of Imathia, but only at less than 3% of the total surveyed rotted peach fruit and blighted shoots. The pathogen overwintered as sub-epidermal pycnidia in blighted shoots or mummified fruit that remained on peach trees. This study also showed that the optimum temperature for mycelial growth and conidial germination of N. parvum was 25 oC. Pathogenicity tests using peach fruit showed that isolates of N. parvum and Diplodia seriata (isolated from pistachio grown in the same region) showed no significant differences in their virulence. In laboratory inoculation tests using detached shoots from 25 peach and nectarine cultivars, N. parvum isolates obtained from rotted peaches caused different size cankers on these cultivars. The cultivar Big Top was the most susceptible while the cultivar Maria Bianca the least susceptible.     

Effects of fruit maturity and wetness on the infection of apple fruit by Neonectria galligena

Neonectria galligena can cause European canker of apple as well as fruit rot. Healthy unwounded fruits on potted trees of cvs Cox, Bramley and Gala were inoculated with conidia of N. galligena to investigate the effects of wetness duration and fruit maturity on rot development. Overall, the incidence of fruit rot was influenced more by fruit maturity at the time of inoculation than by duration of wetness (6–48 h). Young fruit were most susceptible to infection, with 50% of fruit infected when inoculated up to 4 weeks after full bloom. The susceptibility decreased initially until c. 2 months after full bloom and then increased gradually until harvest. Almost all preharvest symptoms (eye rot) developed only on the fruit inoculated up to 4 weeks after full bloom. All other rots were observed after six‐month postharvest storage under controlled atmospheric conditions. However, the relative proportion of preharvest eye rots and postharvest storage rots varied greatly among three years. The effect of wetness duration was only significant for fruit inoculated in their early stages of development but not for those inoculated near harvest. Regression models were developed to describe the observed effects of fruit maturity and wetness on the incidence of total nectria rots.     

Construction of a recombinant strain of <Emphasis Type="Italic">Pseudomonas fluorescens</Emphasis> producing both phenazine-1-carboxylic acid and cyclic lipopeptide for the biocontrol of take-all disease of wheat

Four Neofabraea species are responsible for bull’s eye rot, which is an important postharvest disease of apples and pears. The species diversity of its causal agents in Europe has not been thoroughly explored using molecular genetic methods. Eighty-one Neofabraea isolates were obtained mostly from apples with bull’s eye rot symptoms in the Czech Republic over a two year period. The isolates were identified using PCR fingerprinting and DNA sequencing of the ITS rDNA region, the mitochondrial SSU rDNA and the β-tubulin and EF1α genes. The most common species was N. alba (89 %), followed by N. perennans (5 %) and N. kienholzii (5 %). This is the third published record of N. kienholzii in Europe. The species identity of the isolate CPPF507, which was placed close to N. kienholzii, remains unclear. EF1α was shown to be a suitable marker for the identification of species of the genus Neofabraea and was comparable to the previously used β-tubulin gene. Furthermore, the aggressiveness of individual species was compared and species distribution across Europe was summarized. N. perennans and isolate CPPF507 proved to be the most aggressive, whereas the least aggressive was N. kienholzii. Two N. alba isolates isolated from symptomless apple fruits and leaves were pathogenic to apples in the infection tests.     

Identification of Neofabraea species causing bull's eye rot of apple in Poland and their direct detection in apple fruit using multiplex PCR

Based on partial sequence analysis of the β‐tubulin gene, 19 isolates of fungi causing bull's eye rot on apple in Poland were classified into species: Neofabraea alba, N. perennans and N. kienholzii. To the authors’ knowledge, the detection of N. kienholzii is the second in Europe and the first in Poland. Species affiliation of these fungi was confirmed by a new species‐specific multiplex PCR assay developed on the basis of previously published methods. The new protocol allowed for the specific identification of bull's eye rot‐causing species, both from pure cultures and directly from the skin of diseased or apparently healthy apples. In 550 samples of diseased fruits collected from nine cold storage rooms located in three regions of Poland, in 2011 and 2012, N. alba was detected as the predominant species causing bull's eye rot, occurring on average in 94% of the tested samples. Neofabraea perennans was found in a minority of apple samples, N. kienholzii was found only in two apple samples, while N. malicorticis was not detected in any sample tested. In tests on 120 apparently healthy fruits, only N. perennans was detected in a single sample. The results of genetic diversity analyses of bull's eye rot‐causing fungi based on the β‐tubulin gene sequence and an ISSR (inter‐simple sequence repeat) PCR assay with two primers were consistent, showing the expected segregation of tested isolates with respect to their species boundaries. However, the genetic distance between N. perennans and N. malicorticis was very low, as reported previously.     

Monilia mumecola, a new brown rot fungus on Prunus mume in Japan

In 1982, an anamorphic fungus in the genus Monilia was first isolated as the causal agent of brown rot disease of Japanese apricot or mume (Prunus mume) in Oita Prefecture, Kyushu, Japan. Inoculation of flowers, shoots, and fruit of P. mume with the fungus reproduced brown rot disease symptoms similar to those found in nature. The fungus somewhat resembled the colony appearance of Monilinia (anamorph Monilia) laxa, the apricot brown rot fungus, on PSA plates, but it differed from the latter and the other two brown rot fungi, M. fructigena and M. fructicola, in terms of growth rate, temperature optima for mycelial growth and sporulation, morphology and germination pattern of conidia, nuclear number in the conidium, and nucleotide sequences in the ITS region of ribosomal DNA. It is newly described as Monilia mumecola Y. Harada, Y. Sasaki & T. Sano. A key to anamorphic states of four brown rot fungi of fruit trees is provided.     

Comparative spatial analysis of the sooty blotch/flyspeck disease complex,bull’s eye and bitter rots of apples

The incidence of sooty blotch/flyspeck (SBFS) and bitter and bull’s eye rots were assessed in a Fuji apple orchard during two seasons. Using a regular sampling design, 252 trees were selected and 20 fruits per tree were sampled at harvest and scored for disease incidence. For bitter and bull’s eye rots, additional assessments were made on all symptomless fruit after a 30‐day period of storage. Randomness in the spatial pattern was assessed using beta‐binomial analysis of incidence data for three sampling scales (one, three or six adjacent trees as sampling units) and using Spatial Analysis by Distance Indices (sadie ) for disease counts for the 3‐tree sampling scale. sadie was also used for testing spatial associations between a pair of diseases, between years for the same disease or between rotted and latently infected fruit. Using a toroidal‐shifts procedure, 360 maps of disease counts were created based on the observed data, which were further analysed using sadie . Most datasets showed an aggregated spatial pattern, which was more consistent for the two fruit rots than SBFS, which showed distinct patterns depending on the year or method of analysis. The two fruit rots were spatially associated in most situations but SBFS and bull’s eye rot were dissociated in one season. Results from virtual orchards showed that the patterns observed in the original maps may accurately represent those in similar apple‐growing areas. Hypotheses regarding aspects of ecology and epidemiology of pathogens studied and potential efficacy of control measures in the region are discussed.     

Spatiotemporal analyses of rhizopus rot progress in peach fruit inoculated with Rhizopus stolonifer

Rhizopus rot, caused by Rhizopus stolonifer, is one of the main postharvest diseases in stone fruits, but there is little known about the processes of disease development during transport and postharvest storage. The objective of this study was to characterize temporal progress and spatial distribution of the disease in peach fruit. Rhizopus rot development was evaluated using two different fruit arrangements. Only one fruit of each arrangement was inoculated with a R. stolonifer spore suspension. Disease incidence and severity were assessed daily for all the fruit. Nonlinear models were fitted to the quantity of fruit and to the area of fruit that became infected over time and distance in relation to the source of inoculum. Disease‐free fruit placed next to the artificially inoculated peaches showed disease symptoms due to pathogen dissemination by mycelial stolons. The disease incidence and severity progress rates varied from 0.33 to 0.53 day^?1 and from 0.30 to 0.49 day^?1, respectively. The spatial spread of the disease followed a dispersive wave pattern with increasing speed over time, but decreasing speed with disease severity. For disease severity y = 0.5, the velocity at day 3 varied from 0.14 to 0.32 fruit diameter day^?1, while it ranged from 0.38 to 1.46 fruit diameter day^?1 at day 12.     

RETRACTED ARTICLE: Effect of calcium chloride on the biocontrol efficacy of two antagonistic yeasts against <Emphasis Type="Italic">Penicillium expansum</Emphasis> on apple fruit

Two antagonistic yeasts, Candida membranaefaciens and Pichia guilliermondii, were evaluated for the control of the blue mold of apple caused by Penicillium expansum. Dual culture, cell-free metabolite and volatile tests were used for in vitro assay. Yeast strains of two genera inhibited growth of P. expansum; inhibition varied from 30.27% to 44.19% in dual culture, from 79.40% to 90.57% in the volatile metabolite test, and from 72.99% to 88.77% in the cell-free metabolite test. Calcium chloride (2% w/v) significantly inhibited the growth of the pathogen P. expansum, but did not affect the colony-forming units (CFU) of the yeasts C. membranaefaciens and P. guilliermondii in potato dextrose broth. The concentration of yeast suspension influenced spore germination and germ tube growth of P. expansum in vitro, as well as disease incidence and lesion development in fruits. There were significant negative relationships between the suspension concentrations of the yeasts and the growth as well as infectivity of the pathogen. The addition of calcium resulted in lower spore germination rates and slower growth of germ tubes in vitro, as well as in lower disease incidences and smaller lesion diameters compared with treatments with yeast antagonists alone. When yeast cell suspensions reached a concentration of 10⁷ CFU ml^-1, growth of the pathogen was completely limited in vitro, and no infection was found in apple fruits treated with or without calcium. This article has been retracted because part of the data shown has already been published before, by different authors. An erratum to this article can be found at     

Inhibition on brown rot disease and induction of defence response in harvested peach fruit by nitric oxide solution

Nitric oxide (NO) is an important signal molecule involved in numerous plant responses to biotic and abiotic stresses. The effect of nitric oxide (NO) solution on pathogen infection and defence response of peach (Prunus persica (L.) Batsch) fruit against brown rot disease caused by Monilinia fructicola was investigated. The results showed that 15 μmol l^?1 NO solution did not significantly inhibit spore germination, germ tube length or pathogenicity of M. fructicola, but significantly reduced disease incidence and lesion areas in the fruit. Although 100 μmol l^?1 NO solution effectively inhibited the spore germination, germ tube elongation and pathogenicity of M. fructicola, the high concentration of NO solution caused damage to the fruit. Moreover, 15 μmol l^?1 NO enhanced the activities of chitinase (CHI) and β-1,3-glucanase (GNS) in the fruit. RT-PCR analysis showed that the expression of four genes, CHI, GNS, pathogenesis-related protein 1 and 10 genes (PR-1, PR-10) all increased after NO treatment. Conversely, pretreatment with 100 μmol l^?1 NO scavenger, 2-4-carboxyphenyl-4,4,5,5- tetramethylimidazoline-1-oxyl-3-oxide (cPTIO), rendered the fruit relatively susceptible to pathogen infection and inhibited the defence response of the fruit. These results suggest that NO solution treatment can protect peach fruit from pathogen infection by inducing the activities of the defence enzymes and the expression of PR genes.     

Fusarium rot of melon is caused by several Fusarium species

Fusarium rot of melon, caused by species of the genus Fusarium, has become an important postharvest disease for many Brazilian producers. Due to the delayed onset of symptoms, this disease is often only detected when fruits arrive at the importing country, thus generating economic loss for the exportation of the fruit. This study was developed with the aim of investigating which Fusarium species cause fruit rot in melon and to evaluate any differences in aggressiveness and development of symptoms. Species were identified through phylogenetic analysis of two loci and morphological markers. The 28 isolates obtained from diseased melon fruits of different commercial cultivars were identified as Fusarium falciforme (FSSC), F. sulawesiense, F. pernambucanum (FIESC), and F. kalimantanense (FOSC). Three isolates belong to a new phylogenetic lineage within the F. fujikuroi species complex (FFSC). All isolates were tested for pathogenicity, and first symptoms of rot in Canary melon were observed 2 days after inoculation. Isolates of F. falciforme and F. sulawesiense were shown to be more aggressive. Our results extend information on Fusarium species that cause fruit rot in melon and support the development of management strategies, as there is currently no efficient control for this disease. To our knowledge, this is the first report of the occurrence of species of the FSSC, FOSC, and FFSC from muskmelon fruits in Brazil.     

Volatile organic compounds (VOCs) from cereal plants infested with crown rot: their identity and their capacity for inducing production of VOCs in uninfested plants

When infested with Fusarium sp., the cereals Triticum aestivum L. emend. Fiori et Paol. cv. ‘Bombona’, Avena sativa L. cv. ‘Deresz’, and Hordeum vulgare L. cv. ‘Rastik’ can emit volatile organic compounds (VOCs). The VOCs differ both qualitatively and quantitatively from those emitted by non-infested wheat, oat, and barley plants. We detected increased amounts of VOCs released by green leaves (green leaf volatiles – GLVs): (Z)-3-hexenal, (E)-2-hexenal, (Z)-3-hexen-1-ol, (E)-2-hexen-1-ol, (Z)-3-hexen-1-yl acetate, 1-hexyl acetate as well as the other VOCs like (Z)-ocimen, linalol, linaloloxide, benzyl acetate, indole, and β-caryophyllene. The lipoxygenase pathway resulted in the highest release of GLVs in comparison to the other biochemical pathways of volatile production. As a result of Fusarium infestation, the amounts of VOCs varied between tested cereals. We also subjected uninfested wheat, barley, and oat plants to infested wheat plants, and found that these cereals released larger amounts of VOCs compared to control plants. Emitted amounts of VOCs were significantly higher the shorter the distance between uninfested and infested plants.     

Distribution and virulence of fungal species isolated from yam (Dioscorea spp.) tubers in three agroecological zones of Nigeria

Abstract

Field surveys were conducted in 2012 and 2013 to evaluate the distribution and virulence of fungal isolates associated with tuber rot disease among yam (Dioscorea spp.) producing agroecological zones (AEZs) of Nigeria. A total of 70 and 66 farmers’ fields were surveyed in 2012 and 2013, respectively among three yam species, Dioscorea rotundata, D. alata and D. cayenensis. Fifteen rotted samples were randomly collected per each yam species in fourteen districts across the humid rainforest (HF), the derived savanna (DS) and the southern Guinea savanna (SGS) agroecologies. Twenty-one fungal species were isolated in the HF, 18 species in the DS and 16 in the SGS. The highest rot severity was in HF agroecology in the D. rotundata which varied from 27.2% to 65.7% in 2012 and from 21.1 to 57.4% in 2013, while the SGS agroecology had the least severity. Lasiodiplodia theobromae was the most virulent pathogen with lesion diameter of 26.8–42.1?mm at 14?days after inoculation. Findings provide baseline information for the formulation of management strategies for yam tuber rot disease in Nigeria.     

马铃薯糖苷生物碱对枸杞镰孢菌果腐病的诱导抗性及相关防御酶活性的影响

为探索马铃薯糖苷生物碱诱导采后枸杞鲜果抗病性的效应,采用菌丝生长速率法测定了离体条件下马铃薯糖苷生物碱对枸杞鲜果采后主要致腐病原菌镰孢菌Fusarium sp.的抑菌活性;采用浸泡处理法测定了马铃薯糖苷生物碱对采后枸杞鲜果发病病情指数及对果实多酚氧化酶(PPO)、过氧化物酶(POD)、苯丙氨酸解氨酶(PAL)和超氧化物歧化酶(SOD)等相关防御酶活性的影响。结果表明:不同浓度马铃薯糖苷生物碱对镰孢菌均具有一定的抑制作用,EC50为0.11 g/mL;0.05～0.25 g/mL(提取原料)的马铃薯糖苷生物碱能显著降低采后枸杞鲜果病情指数,较对照降低了10.18%～38.51%;以0.15 g/mL(提取原料)的马铃薯糖苷生物碱处理后对枸杞鲜果抗果腐病的诱导效果最好,达45.81%;马铃薯糖苷生物碱处理枸杞鲜果后,果实中4种防御酶PPO、POD、PAL、SOD的活性均较对照有不同程度提高,分别于处理后第4、5、2、2天与对照差异达到最大,较对照提高了30.76%、21.34%、31.35%和21.91%。表明马铃薯糖苷生物碱能够诱导枸杞鲜果对采后病害的抗性效应,且采后抗病性可能与枸杞相关防御酶活性的增加有关。     

Characterization of Glomerella cingulata isolates from almond fruit using VCG,molecular, fungicide sensitivity and pathogenicity tests 1

Almond anthracnose caused by Glomerella cingulata is a major disease of this crop in Israel. The pathogen infects young fruit resulting in fruit rot. Leaf wilting and shoot dieback accompany fruit rot, even though the pathogen cannot be isolated from leaves or twigs. Isolates of G. cingulata from diseased almond fruit were compared using vegetative compatibility grouping (VCG), molecular methods, fungicide sensitivity and pathogenicity assays in order to determine the genetic diversity and host specificity among different populations. Polymerase chain reaction amplification of genomic DNA, using four primers produced uniform banding patterns for all the almond isolates from different geographic locations in Israel. HaeIII digestion patterns of A + T-rich DNA, and Southern hybridization of the repetitive nuclear DNA element (GcpR1) to PstI-digested genomic DNA of almond isolates also revealed no polymorphism. Chlorate-resistant nitrate-nonutilizing (nit) mutants were generated and used in heterokaryon tests. Complementary heterokaryons formed between the mutants of different isolates indicated a single VCG. Isolates of G. cingulata from almond had optimal growth temperatures of 20–22°C as opposed to 26–28°C for avocado isolates. In addition, almond isolates of G. cingulata are insensitive to benzimidazole fungicides in contrast to sensitivity of isolates from avocado. In artificial inoculations, almond isolates infected almond, avocado, apple, mango and nectarine fruit at a slower rate than G. cingulata isolates from avocado, apple and mango. Only the anamorph Colletotrichum gloeosporioides has been detected on almond in Israel, whereas isolates of G. cingulata from other hosts produce ascocarps.     

Phenazines are Involved in Biocontrol of Pythium myriotylum on Cocoyam by Pseudomonas aeruginosa PNA1

Root rot of cocoyam (Xanthosoma sagittifolium) caused by Pythium myriotylum is the most devastating disease of this important tropical tuber crop with yield reductions of up to 90%. Bioassays were conducted in vitro and in sterile volcanic soil artificially infested with Pythium myriotylum, isolate CRPm, to test whether Pseudomonas aeruginosa PNA1 can control the cocoyam root rot disease. P. aeruginosa PNA1 (wild type) produces phenazine-1-carboxylic acid and phenazine-1-carboxamide (oxychlororaphin), while its tryptophan auxotrophic mutant FM13 is phenazine negative and secretes anthranilate in vitro. PNA1 and FM13 have previously been shown to control Pythium debaryanum and Pythium splendens on lettuce and bean. PNA1 and FM13 significantly inhibited growth of P. myriotylum in dual cultures, while their supernatants highly reduced mycelial dry weight in potato dextrose broth. However, in the presence of tissue culture derived cocoyam plantlets, only strain PNA1 strongly reduced root rot disease severity. Soil experiments involving strain PNA1 in comparison to phenazine-deficient mutants suggested that the biocontrol activity of PNA1 against P. myriotylum may involve phenazines. Phenazine involvement was further strengthened by the fact that FM13 fed with exogenous tryptophan (so that phenazine production is restored) significantly reduced disease severity on cocoyam. The efficiency of PNA1 to control P. myriotylum on cocoyam was significantly improved when the strain and the pathogen were allowed to interact for 24 h prior to transplanting cocoyam plantlets, while doubling the inoculum density of the pathogen negatively affected its efficiency.