Screening and identification of yeast strains from fruits and vegetables: Potential for biological control of postharvest chilli anthracnose (Colletotrichum capsici)

Yeasts antagonistic to Colletotrichum capsici were isolated from Thai fruits and vegetables. Four antagonists (R13, R6, ER1, and L2) were found that inhibited C. capsici growth with biocontrol efficacies of 93.3%, 83.1%, 76.6%, and 66.4%, respectively. Identification by 26S rDNA, and ITS region sequence together with physiological and morphological characteristics, showed them to be Pichia guilliermondii, Candida musae, Issatchenkia orientalis, and Candida quercitrusa, in order of their efficacy. P. guilliermondii strain R13 showed efficacy in reducing disease incidence on C. capsici infected chilli fruits to as low as 6.5%. Lower disease incidence was observed at lower storage temperature. The application of P. guilliermondii is more effective for preserving chilli fruits than conventional preservation with chlorinated water.     

Identification and Characterization of two new Fungal Pathogens of Polygonatum odoratum (Angular Solomon's seal) in Italy

Polygonatum odoratum (Mill.) Druce (Angular Solomon's seal) is a well‐known ornamental plant and herbal drug, which is used in traditional medicine. Severe leaf spots were observed in wild Angular Solomon's seal plants in a public park in Trento, Italy. A combination of morphological and molecular characteristics, including sequence data of ITS‐rDNA, large subunit (LSU), beta tubulin (TUB) and glyceraldehyde 3‐phosphate dehydrogenase (GAPDH) led to identification of two species, namely Phoma odoratissimi and Colletotrichum dematium s. str., isolated from these leaf spots. Pathogenicity of the isolates was confirmed on wounded leaves of Po. odoratum; indeed, both species induced leaf spot symptoms on inoculated leaves within 10 days of inoculation. This is the first report of leaf spot disease caused by Ph. odoratissimi and C. dematium s. str. on Po. odoratum, which can be considered a new host for both the species examined in this study. In addition, isolation of Ph. odoratissimi represents a new record for the mycobiota of Italy.     

Adjuvants enhance the biological control potential of an isolate of Colletotrichum gloeosporioides for biological control of sicklepod (Senna obtusifolia)

In greenhouse experiments, unrefined corn oil, Silwet L-77, and an invert emulsion were tested as adjuvants with the mycoherbicidal fungus Colletotrichum gloeosporioides, a weakly virulent pathogen of sicklepod (Senna obtusifolia). A 1:1 (v/v) fungus/corn oil tank mixture containing 0.2% (v/v) Silwet L-77 surfactant reduced the dew period requirements for maximum weed infection and mortality from 16 to 8 h, and delayed the need for free moisture for greater than 48 h. This formulation also resulted in the ability of the pathogen to infect and kill weeds in larger (>5 leaf) growth stages. The invert emulsion resulted in similar effects upon these parameters. These results suggest that invert emulsions, unrefined corn oil and Silwet L-77 surfactant greatly improve the bioherbicidal potential of this pathogen for control of sicklepod, a serious weed pest in the southeastern US.     

Set‐point control of RD21 protease activity by AtSerpin1 controls cell death in Arabidopsis

Programmed cell death (PCD) in plants plays a key role in defense response and is promoted by the release of compartmentalized proteases to the cytoplasm. Yet the exact identity and control of these proteases is poorly understood. Serpins are an important group of proteins that uniquely curb the activity of proteases by irreversible inhibition; however, their role in plants remains obscure. Here we show that during cell death the Arabidopsis serpin protease inhibitor, AtSerpin1, exhibits a pro‐survival function by inhibiting its target pro‐death protease, RD21. AtSerpin1 accumulates in the cytoplasm and RD21 accumulates in the vacuole and in endoplasmic reticulum bodies. Elicitors of cell death, including the salicylic acid agonist benzothiadiazole and the fungal toxin oxalic acid, stimulated changes in vacuole permeability as measured by the changes in the distribution of marker dye. Concomitantly, a covalent AtSerpin1–RD21 complex was detected indicative of a change in protease compartmentalization. Furthermore, mutant plants lacking RD21 or plants with AtSerpin1 over‐expression exhibited significantly less elicitor‐stimulated PCD than plants lacking AtSerpin1. The necrotrophic fungi Botrytis cinerea and Sclerotina sclerotiorum secrete oxalic acid as a toxin that stimulates cell death. Consistent with a pro‐death function for RD21 protease, the growth of these necrotrophs was compromised in plants lacking RD21 but accelerated in plants lacking AtSerpin1. The results indicate that AtSerpin1 controls the pro‐death function of compartmentalized protease RD21 by determining a set‐point for its activity and limiting the damage induced during cell death.     

Differential Organ Distribution,Pathogenicity and Benomyl Sensitivity of Colletotrichum spp. from Blackberry Plants in Northern Colombia

Blackberry anthracnose, caused by Colletotrichum spp., is an important disease of cultivated blackberry in the world. In Colombia, it is the number one limiting factor for commercial production. This study was conducted to determine the species of Colletotrichum infecting blackberry plants as well as the organ distribution, pathogenicity and response to benomyl of the isolated strains. Sixty isolates from stems (n = 20), thorns (n = 20) and inflorescences (n = 20) were identified as Colletotrichum acutatum and Colletotrichum gloeosporioides by a species‐specific polymerase chain reaction (PCR). Both Colletotrichum species were found in the same plant but on different organs. Colletotrichum gloeosporioides species predominated in thorn lesions (n = 16) and C. acutatum in stems (n = 15) and inflorescence (n = 15). Pathogenicity assays on detached blackberry organs demonstrated differences between the two species with an average period of lesion development of 8.7 days for C. gloeosporioides and 10.3 days for C. acutatum. Wound inoculated organs had 90% disease development compared to 17.5% in non‐wounded. All C. acutatum isolates (n = 34) were benomyl tolerant, whereas C. gloeosporioides isolates (n = 26) were 30.7% sensitive and 69.2% moderately tolerant. Phylogenetic analysis with ITS sequences of a subset of 18 strains showed that strains classified as C. gloeosporioides had 100% identity to Colletotrichum kahawae, which belongs to the C. gloeosporioides species complex, whereas C. acutatum strains clustered into two different groups, with high similarity to the A2 and the A4 molecular groups. These data demonstrate for the first time the differential distribution of both species complexes in blackberry plant organs and further clarifies the taxonomy of the strains.     

Silicon and Fungicide Effects on Anthracnose in Moderately Resistant and Susceptible Sorghum Lines

This study aimed to evaluate the effect of silicon (Si) and its interaction with fungicide on the management of sorghum anthracnose. The experiments were carried out in Si‐deficient soil in the 2008/2009 and 2009/2010 growing seasons in a randomized, complete block, split‐split plot design with four replications. Calcium silicate (CS) and lime (L), at the rates of 6 and 5 ton/ha, respectively, were randomly assigned to the main plot. Two sorghum lines, BR‐008 (resistant) and BR‐009 (susceptible), were assigned to the split plots. The split‐split plots corresponded to with or without the fungicide Opera® (epoxiconazole + pyraclostrobin). The residual effect of CS and L from the 2008/2009 growing season was evaluated in the 2009/2010 growing season. For the 2008/2009 growing season, the area under anthracnose progress curve (AUAPC) was reduced by 39 and 42% for lines BR‐008 and BR‐009, respectively, with the application of CS. In the presence of the fungicide, the AUAPC was reduced by 35 and 42% for the CS and L treatments, respectively. Calcium silicate with and without fungicide contributed to decreasing the AUAPC by 44 and 37%, respectively. The fungicide spray decreased the AUAPC by 50 and 39% for lines BR‐008 and BR‐009, respectively. Without fungicide, the AUAPC decreased by 88% for line BR‐008 compared with line BR‐009; however, with fungicide, the reduction reached 90%. The Si leaf tissue concentration significantly increased with the CS application (5.9 g/kg) compared with the L application (0.3 g/kg), regardless of the sorghum line. The yield increased by 0.6 ton/ha with the CS compared to the L application. The fungicide increased yield by 0.48 ton/ha compared with the non‐fungicide spray treatment. The residual effect of CS in the soil increased Si leaf tissue concentration and yield as well as reduced the intensity of anthracnose in the 2009/2010 growing season.     

Fungicidal Properties of Some Novel Trifluoromethylphenyl Amides

Trifluoromethylphenyl amides (TFMPAs) were designed and synthesized as potential pesticides. Thirty‐three structures were evaluated for fungicidal activity against three Colletotrichum species using direct bioautography assays. Active compounds were subsequently tested against C. fragariae, C. gloeosporioides, C. acutatum, Phomopsis obscurans, P. viticola, Botrytis cinerea and Fusarium oxysporum. The study identified 2‐chloro‐N‐[2,6‐dichloro‐4‐(trifluoromethyl)phenyl]acetamide ( 7a ) as showing the strongest antifungal activity, and the broadest activity spectrum in this set against Colletotrichum acutatum (at 48 and 72 h) and Phomopsis viticola (at 144 h). The presence of triethylamine in its complex with N‐[2,6‐dichloro‐4‐(trifluoromethyl)phenyl]‐2,2,3,3,3‐pentafluoropropanamide ( 7b′ ) played an important role in the bioactivity, and depending on the concentration or fungal species it showed higher or lower activity than the parent amide. X‐Ray crystallography has shown that the complex ( 7b′ ) is an ion pair, (C₁₀H₂Cl₂F₈NO)^? (C₆H₁₆N)⁺, where a proton is transferred from the amide nitrogen to the triethylamine nitrogen and then connected by hydrogen bonding to the acyl oxygen (N?H 0.893 Å; H???O 1.850 Å; N???O 2.711 Å; N?H???O 161.2(13)°). Although none of these compounds were better than standards, this work revealed some potential lead structures for further development of active novel compounds.     

In vitro screening of biological and chemical agents together on the growth of Colletotrichum gloeosporioides (Penz.) Penz. and Sacc. causing inflorescence die back in arecanut

Colletotrichum gloeosporioides is a destructive pathogen of many crop species causing diseases in many annual, biennial and perennial plants. A study was undertaken to find out the effect of biological and chemical agents together on the growth of C. gloeosporioides causing inflorescence die back in arecanut at the Department of Plant Pathology, CPCRI, Kasaragod. To reduce the release of chemical pesticides to the environment, integrated control strategies have been adopted extensively by combining both bioagents and chemical agents. So in the present study in vitro experiments were conducted with two compatible Trichoderma sp. viz., Trichoderma virens and Trichoderma viride and fungicides viz. Blitox 50 W and Mixol 72. The results indicated that all the treatments had a significant inhibitory effect on the growth of C. gloeosporioides and reduced its colony diameter. High percent inhibition was found when 0.05% of Mixol 72 was used with T. virens (87.61%). The least inhibition was shown by T. virens+0.05% Blitox 50 W (80.95%). It is concluded that the combination of bioagents with fungicides provided higher disease suppression than achieved with fungicides and bioagents when used alone.     

Detection of Colletotrichum falcatum causing red rot of sugarcane by enzyme-linked immunosorbent assay

Red rot disease of sugarcane caused by Colletotrichum falcatum Went is one of the most destructive diseases of sugarcane (Saccharum officinarum L.) worldwide. The pathogen spreads primarily through infected sugarcane setts and hence the use of disease-free setts is essential to prevent the disease. In order to develop immunological method for detection of C. falcatum, two proteins with molecular weights of 27 kDa and 45 kDa were purified from the mycelium of C. falcatum race Cf 05 and used as antigen source to raise polyclonal antibodies in NewZealand white rabbit. The developed polyclonal antibodies were tested for detection of C. falcatum by enzyme-linked immunosorbent assay (ELISA) and immunoblot analysis. The polyclonal antibodies specifically detected C. falcatum in extracts from infected plants, both in immunoblot and ELISA. The ELISA results showed that the developed polyclonal antibodies were highly specific to C.falcatum. The developed antibodies were very sensitive and could detect C.falcatum proteins even at a dilution of 1:50,000. Higher ELISA absorbance values were recorded even at an antigen dilution of 1:500. In western blot analysis, protein bands with molecular weights of 27 kDa and 45 kDa reacting to antisera raised against 27 kDa and 45 kDa mycelial proteins of C. falcatum, respectively, were detected in protein samples from red rot infected canes. The high specific reactivity and sensitivity of the antisera indicate its potential suitability for ELISA-based detection of C. falcatum.