Banana Anthracnose Research Articles

Zein nanoencapsulation enhances the antifungal activity of thymol for postharvest decay control in bananas

AbstractThe current work describes a nanoparticle system-based approach to enhance the antifungal activity of thymol, a ubiquitous natural antifungal phenolic compound, in postharvest control against banana anthracnose. Thymol was encapsulated within the amphiphilic protein zein by high-shear emulsification, yielding highly dispersible thymol-loaded zein nanoparticles with a high encapsulation efficiency (70%). These particles have an average diameter of 300 nm with spherical morphology, smooth interface, and matrix-type internal structure, as supported by comprehensive structural characterization (dynamic light scattering, transmission electron microscopy and atomic force microscopy). Based on a 40-d storage stability test, thymol was effectively retained within the nanoparticles at 4 °C and ambient room temperature (99% and 97% retention, respectively), despite thymol’s instability and volatility. Antifungal activity assessment against Colletotrichum musae, one of the predominant pathogens that cause banana anthracnose, showed a 200- to 300-fold improvement in the in vitro antifungal activity of thymol. Moreover, the application of thymol-loaded zein nanoparticles as a spray component for banana postharvest treatment demonstrated the efficacy of thymol-loaded zein nanoparticles in preventing and delaying the formation of initial symptoms of banana anthracnose. This appears to arise from the thymol-loaded zein nanoparticles depositing as a film on the banana epidermis, as revealed by atomic force microscopy. Overall, this nanoparticle system offers a new avenue for the design of effective antifungal materials with potential applications in combatting postharvest diseases. Graphical abstract

Solid Formulation of <em>Trichoderma virens</em> for the Management of Banana Anthracnose Caused by<em> Colletotrichum musae</em>

Trichoderma spp. widely used as biocontrol agents for controllin g a wide range of plant diseases. Banana anthracnose caused by Colletotrichum musae is a prominent, widely distributed postharvest disease. This study was carried ou t to assess the effectiveness of locally isolated Trichoderma virens against C. musae and the mass producti on of T. virens using locally available, low-cost solid substrates. T. virens was tested for the inhibition of C. musae isolate in -vitro and in-vivo by dual plating and inoculating into two varieties of ripened banana: Kolikuttu and Cavendish, respectively. For mass production, T. virens was inoculated into different solid substrates including scraped coconut waste, sawdust, tea waste, seeds from rice, finger millet, and maize, dried pieces of water hyacinth plant, paddy straw, and Panicum maximum leaves; regularly taken spore co unts (cfu/g) and checked for viability by plating after 12 weeks of storage. The pathogen inhibition percentag e by T. virens was 74.10%. Disease severity was 0% in Kolikuttu and 19% in Cavendish after 5 days of T. virens spore appli cation (1x107 spores/ml). Significantly higher (p≤0.05) mean spore production resulted in rice seeds (9.345x109 spores/g) compared to the other substrates and the least resulted in sawdust (1.808x109 spores/g) at the 8th week after T. virens inoculation. Spores of T. virens were viable in all the tested substrates throughout the study period. The results conclude that T. virens is capable of controlling banana anthracnose and can be efficiently mass-produced by using rice seeds, dried pieces of P. maximum leaves, and f inger millet seeds as substrates.

Comparative Evaluation of Different Biocontrol Agents against Anthracnose of Nendran Banana Caused by Colletotrichum musae

India is the largest producer of banana in the world. Nendran, a French plantain, is the most important banana cultivar in Kerala. It is mainly affected by many diseases both in the field and after harvest. Being a freshly consumed fruit crop there are a lot of limitations in using chemicals for its disease control. Keeping this view, a study was conducted at the Department of Plant Pathology, College of Agriculture, Vellayani, to control anthracnose, a serious post harvest disease under tropical conditions. In this study comparative evaluation of three fungal biocontrol agents namely [Trichoderma sp. (KAU), Trichoderma sp. (TRPN-3), Trichoderma sp. (TRKR-2)] and bacterial biocontrol agents [Pseudomonas fluorescens (PN026), Bacillus sp. (B15), Bacillus sp. (B17)] were tested against Colletotrichum musae. Among the fungal biocontrol agents maximum mycelial growth inhibition was recorded with Trichoderma sp. (KAU) (77.44%) followed by Trichoderma sp. (TRKR-2) (77.11%). Minimum mycelial growth inhibition was recorded in P.fluorescens (PN026) (55.33%). Based on the results reveals that Trichoderma sp. (KAU) is a highly promising biocontrol agent for managing anthracnose of banana.

Biocontrol efficiency and potential mechanism of streptomyces distatochromogenes XT34 against postharvest anthracnose caused by colletotrichum musae on banana fruit

Banana anthracnose caused by Colletotrichum musae is a serious postharvest disease. Use of fungicides has potential adverse effects on the environment safety and human health. Application of biological control agents (BCA) is a promising strategy for managing postharvest fruit diseases. In this study, Streptomyces sp. XT34 was isolated from the banana rhizosphere soil and exhibited a strong antifungal activity against C. musae. Combining the phenotypic and whole-genomic alignment, strain XT34 was identified as Streptomyces diastatochromogenes. Strain XT34 extracts reduced the disease incidence of anthracnose and maintained the fruit quality of banana. Treatment of extracts reduced the colonization of C. musae on fruit surface. The activity inhibition of cell wall-degrading enzymes maintained the cell-wall strength of fruit. The activity increase of defense enzymes contributed to the low disease symptom. It was supported by the differentially expressed genes (DEGs) related to supramolecular fiber organization and flavonoid biosynthesis. In addition, spore germination of C. musae was effectively inhibited by extracts. The morphology of mycelia and spore become wrinkled and ruptured. The treatment of extracts increased the optical density of A260 and soluble sugar and protein contents of C. musae. The integrity of cell membrane was also destroyed, reflecting on an increase in electrical conductivity and malondialdehyde. Several metabolites were found in the genome of strain XT34 using antiSMASH. Eleven volatile compounds were identified by gas chromatography-mass spectrometer (GC-MS). Notably, strain XT34 and extracts exhibited a broad-spectrum antifungal activity against eight phytopathogenic fungi. Hence, S. diastatochromogenes XT34 provides a potential BCA to control banana anthracnose.

Assessment of Various Fungicides, Bioagents and Natural Substances on Anthracnose Disease of Banana

The research was conducted to minimize the activity of harmful microorganism which is responsible for deterioration of fruit quality. The eight treatments were taken during the in-vitro study from which viz., six fungicides and two bioagents were tested for fruit dip treatment against anthracnose of banana. As a result of study, the most effective treatment was found carbendazim 12 % + mancozeb 63 % WP, which is showing the least disease intensity (20.26 %) and the highest reduction in banana anthracnose (52.56 %). Carbendazim and propiconazole also demonstrated significant disease reduction. Among non-hazardous chemicals, chitosan was the most effective (31.11 % disease intensity, 34.24 % reduction), followed by edible paraffin wax and yeast. Hot water treatment showed the highest disease intensity (44.49 %) and the least disease control (20.86 %). The study concluded that, carbendazim 12 %+ mancozeb 63 % WP and chitosan was the most effective against the post-harvest anthracnose disease of banana.

Molecular and phenotypic characterization of <em style="mso-bidi-font-style: normal;">Colletotrichum plurivorum</em> and <em style="mso-bidi-font-style: normal;">Colletotrichum</em> <em style="mso-bidi-font-style: normal;">musae</em> causing banana anthracnose disease in the Central Province of Sri Lanka

Most of the commercial banana cultivars in Sri Lanka are susceptible to anthracnose disease. Colletotrichum musae has been known as the causal agent of banana anthracnose for decades and the pathogen has been identified using morphological characteristics. Molecular analyses based on multigene phylogenetics are now standard protocols to identify Colletotrichum species. The present study was aimed at identifying Colletotrichum species causing banana anthracnose by molecular and phenotypic characterization. Thirty-seven isolates were obtained from ripened bananas showing anthracnose symptoms, collected from different locations in the Central Province of Sri Lanka. Of them, 36 were preliminarily identified as Colletotrichum based on conidial morphology. The remaining isolate did not sporulate during the entire study period. Ten isolates taken for molecular studies consisted of eight with orange/white arial mycelia and orange conidial masses, one with a white to greyish colony and blackish clusters of ascomata, and one with a white to faint pink colour colony. DNA extracted from each isolate was subjected to multi-gene DNA sequence analysis using ITS, TUB, GAPDH and GS loci. Based on phylogenetic analyses, eight isolates were identified as Colletotrichum musae, and the other two as C. plurivorum and C. siamense. The vegetative morphology of C. plurivorum differed considerably from C. musae and C. siamense. Slight differences in colony morphology were observed among the C. musae isolates. Freshly harvested healthy bananas were artificially inoculated with isolates of C. musae or C. plurivorum and produced typical anthracnose lesions within a week. The Colletotrichum siamense isolate failed to develop anthracnose symptoms. This is the first report of C. plurivorum causing banana anthracnose.

A Vis/NIR spectra-based approach for identifying bananas infected with Colletotrichum musae.

Anthracnose of banana caused by Colletotrichum species is one of the most serious post-harvest diseases, which can cause significant yield losses. Clarifying the infection mechanism of the fungi using non-destructive methods is crucial for timely discriminating infected bananas and taking preventive and control measures. This study presented an approach for tracking growth and identifying different infection stages of the C. musae in bananas using Vis/NIR spectroscopy. A total of 330 banana reflectance spectra were collected over ten consecutive days after inoculation, with a sampling rate of 24 h. The four-class and five-class discriminant patterns were designed to examine the capability of NIR spectra in discriminating bananas infected at different levels (control, acceptable, moldy, and highly moldy), and different time at early stage (control and days 1-4). Three traditional feature extraction methods, i.e. PC loading coefficient (PCA), competitive adaptive reweighted sampling (CARS) and successive projections algorithm (SPA), combining with two machine learning methods, i.e. partial least squares discriminant analysis (PLSDA) and support vector machine (SVM), were employed to build discriminant models. One-dimensional convolutional neural network (1D-CNN) without manually extracted feature parameters was also introduced for comparison. The PCA-SVM and·SPA-SVM models had good performance with identification accuracies of 93.98% and 91.57%, 94.47% and 89.47% in validation sets for the four- and five-class patterns, respectively. While the 1D-CNN models performed the best, achieving an accuracy of 95.18% and 97.37% for identifying infected bananas at different levels and time, respectively. These results indicate the feasibility of identifying banana fruit infected with C. musae using Vis/NIR spectra, and the resolution can be accurate to one day.

Management of Banana Anthracnose, Post-Harvest Practices, and Future Aspects – A Review

Management of Banana Anthracnose, Post-Harvest Practices, and Future Aspects – A Review

Essential oil of Piper macedoi Yunck. leaves, potential alternative for the management of banana anthracnose disease

The fungus Colletotrichum musae could cause a disease called anthracnose which reduces the quality of the banana fruit in the postharvest period. Anthracnose causes rots, peel lesions and generate great economic losses. Synthetic fungicides, used as an attempt to reduce the pathogen population, although effective, when used in amounts and frequency above the recommended, cause damage the environment and human health and can induce resistance. Research has focused in the search for alternative methods to reduce the use of synthetic substances in managing of agricultural diseases. The control of pathogens using compounds of natural origin is highlighted among new management techniques. Thus, we aimed to extract and characterize the essential oil from Piper macedoi leaves and evaluate its effect in the inhibiting the mycelial growth of the fungus C. musae in vitro. The essential oil was extracted by hydrodistillation, analyzed and characterized by Gas Chromatography and submitted to assays of fungicidal activity with C. musae. In comparison with dry plant material, yield of P. macedoi essential oil was 0.93 %. Piperitone (26.48 %), silvestrene (13.03 %), and bicyclogermacrene (10.45 %), were the major compounds identified in the essential oil composition considering the percentage in area. Regarding the antifungal activity tests, the essential oil showed a low inhibitory concentration (IC) for the IC50 (0.804 mL.L−1) and IC90 (1.80 mL.L−1), showing that the essential oil from P. macedoi leaves has great potential to control C. musae.

Fungicidal activities of Cymbopogon winterianus against anthracnose of banana caused by Colletotrichum musae

The genus Cymbopogon (Poaceae) species have been widely cultivated throughout the world for a wide range of uses in the pharmaceutical and agricultural fields. The current work investigates the fungicidal activities of Cymbopogon winterianus extract (CWE) in controlling the C. musae that caused anthracnose disease in banana fruit. In vitro assay results showed that CWE at 1.5–2.5 gL−1 concentrations controlled the development of the test pathogen. Mycelial blast, cytoplasmic discharge, and spore edema were noticed when CWE was applied. The Minimum Effective Concentration (MEC) of CWE for the in vivo assay was 1.50 gL−1 and can be used as a postharvest treatment on banana fruit to deter anthracnose infection. Moreover, no visible phytotoxicity or changes in aroma were observed on banana fruit treated with CWE, even at the highest concentration of 2.5 gL−1. The GCMS analysis revealed 41 chemical components associated with CWE. The five main compounds were the following: Methyl oleyl ether (40.20%), γ-Sitosterol (15.80%), 6-Methylheptan-3-ol (7.13%), α-Terpineol (5.56%), and n-Pentadecanol (4.05%). The CWE possesses excellent fungicidal effects against C. musae; in the near future, it can be used as an alternative to commercially available traditional fungicides on the market.

Use of Melaleuca alternifolia essential oil as an efficient strategy to extend the shelf life of banana fruits

Banana anthracnose is one of the main postharvest diseases economically compromising the fruit production chain. Fruit treatment with natural products can reduce postharvest pathogen infestation and increase protection during storage. For natural control of this disease, the activity of the essential oil of Melaleuca alternifolia (MAEO) was investigated in Colletotrichum musae. The aim of this study was to evaluate the in vitro antifungal activity of the MAEO on C. musae by of different concentrations and to evaluate the effect of MAEO on the incidence and severity of anthracnose. The essential oil (EO) was characterized by gas chromatography (GC-FID and GC-MS). The relative percentage of monoterpenes was 96.94%, and terpinen-4-ol was the main component (47.08%). The minimum inhibitory dose (EC100) of the mycelial growth of C. musae was reached at a concentration of 2.15 μg mL−1. Bananas were treated with water emulsions MAEO into at different concentrations (0.50, 1.00, 2.00, 3.00, 4.00 and 5.00 μg mL−1) for 08 days of storage at 25 ± 2 °C. MAEO showed fungistatic action from the concentration of 2.0 μg mL−1 (p < 0.05). The results showed that, after storage, MAEO at 0.50, 1.00, 2.00, 3.00, 4.00 and 5.00 μg mL−1 reduced area under the disease-progress curve (AUDPC) by 80%, 90%, 95%, 96%, 97% and 98%, respectively. MAEO reduces the severity of anthracnose after fruits treatment, without causing phytotoxic action, maybe a promising candidate for Biorational pesticides, in preservation and increment shelf life fruits banana.

Control of banana anthracnose by hot water dip: A semi-empirical model coupling heat transfer and Colletotrichum musae inactivation

A semi-empirical model was developed to predict simultaneously time-temperature distribution inside the unripe banana and fungal reduction during the hot water dip, in order to simulate different process conditions for controlling banana anthracnose disease. For this purpose, the kinetic parameters of Colletotrichum musae reduction were estimated based on the thermal death time concept (z = 3.56 °C and D50°C= 1044 s), and evaluated in different simulated media (unripe and fully ripe banana peel agar media). At the same time, a heat transfer model based on the Fourier equation was developed considering two unripe banana domains (peel and pulp). The comparison between the predicted and observed number of C. musae spores after the thermal treatment of unripe banana was satisfactory (coefficient of variation less than 10%), suggesting that the survival ratio is affected by the banana composition. The resulting model can be used to simulate different time-temperature scenarios, and to show how the parameter combinations may affect C. musae reduction in the unripe banana during the hot water dip process.

In vitro bioassays of some plant extracts against Banana anthracnose caused by Colletotrichum musae

Anthracnose of banana caused by Colletotrichum musae is an invasive disease, which is difficult to control in the export process. Plant extracts used to control plant diseases are gradually becoming substitutes for traditional fungicides because they are safe for the human body and environmentally friendly. The study was conducted to record the prevalence of banana anthracnose in two locations and to determine the best medium for the growth of C. musae and botanical extracts for managing the fungus under in-vitro conditions. Six media viz., Ripen banana peels media (RBPM), Unripen banana peels media (URBPM), Ripen banana flash fruit media (RBFFM), Unripen banana flash fruit media (URBFFM), Banana leaf media (BLM) and Potato dextrose agar media (PDA), four botanical extracts including Garlic (GC), Mint (MT), Eucalyptus (EC) and Neem seed (NS) at 2, 4 and 6% concentrations were tested against the fungus and the control (CL) remain untreated. Potato dextrose agar media (PDA) was used as a standard semi-synthetic growth media. The prevailing frequency of the disease was high in the investigation area. The maximum linear mycelial growth of the C. musae was recorded on PDA (85 mm) followed by URBFFM (40 mm), RBFFM (26 mm) whereas the lowest mycelial growth was recorded on BLM (20 mm) and URBPM (8 mm). In-vitro experiments showed that PDA was the best medium for the mycelial growth of C. musae. Moreover, tested botanical extracts showed high variation in reducing the colony growth of the fungus. EC was found most effective in reducing the colony growth of the fungus followed by NS, GC and MT. These results suggest that all the botanical extracts may be potentially useful for controlling anthracnose of bananas, EC extract can be effectively used for the field experiment. Keywords: Botanical extracts, Efficacy, Growth media, Incidence, Prevalence

Efficacy and potential mechanism of hinokitiol against postharvest anthracnose of banana caused by Colletotrichum musae

Anthracnose caused by Colletotrichum musae is a devastating post-harvest disease on bananas, and it leads to serious economic losses. Exploring environmentally friendly natural compounds to manage anthracnose in fruit has been attracting more attention. In the current study, the effect of hinokitiol against C. musae and on banana quality was investigated. Our results showed that hinokitiol exhibited promising antifungal activities both in vitro and in vivo. Importantly, it effectively delayed banana ripening by maintaining weight, color, firmness, and TSS content, and it dramatically reduced the release of ethylene, the respiration rate, and the expression of genes involved in releasing ethylene. Moreover, the activities of defense-related enzymes including SOD, POD, and APX were enhanced by hinokitiol. The mechanism of hinokitiol against C. musae may be attributed to the destruction on the integrity of the plasma membrane, which leads to the leakage of cellular constituents and eventually the cell death of C. musae. Taken together, this study provided substantial evidence that hinokitiol had great potential to be developed as a natural antifungal agent against C. musae, prolonging the storage period of postharvest bananas.

Antagonistic activity of bacteria from wild honey against Colletotrichum musae, and testing of wild honey as biopesticide spray to control banana anthracnose

Anthracnose is a foliar and fruit disease caused by Colletotrichum spp. affecting a wide range of crops. Infection occurs early followed by quiescence in fruits, such as in banana, where chemical-based pesticides are used as a dependable fungal control for many years. There is an increasing need for a safe control and as implicated in the Organic Agriculture Act of 2010 (RA 10068) in the Philippines. This scenario drove the use of alternative pest control such as the use of biologicals and natural products. In this study, seven bacteria were isolated from wild honey, produced by Apis mellifera, wherein four (BC2, BC3, BC6 and BC7) were found to be an effective antagonist against Colletotrichum musae in in vitro conditions. These bacteria were identified to belong to the genus Lactobacillus spp. (BC2, BC3, BC7) and Bacillus spp. (BC6) based on sugar utilization tests, morphological and cultural growth in PDPA. For the in vivo test, different dilutions of wild honey were used and it was found out that lower concentrations were effective as biopesticide spray to prevent anthracnose infection. Lastly, we report herewith the first isolation of bacteria with biological control potential from wild honey, and to apply the raw or natural product as biopesticide in postharvest fruits.

Mycofumigation with the endophytic fungi Fusarium proliferatum (Matsushima) Nirenberg and Diaporthe sp. for the control of banana and mango anthracnose

Banana and mango are the top commodities in the Philippines that are traded worldwide however, fungal diseases such as anthracnose affect their yield and quality. Environmentally sound control strategies have been explored and one of these is mycofumigation using fungal endophytes. This alternative approach is understudied in the Philippines, hence, the study aimed to evaluate the mycofumigation potential of fungal endophytes collected from Mt. Makiling, Luzon, Philippines and their pathogenicity to banana and mango fruits. In vitro and in vivo mycofumigation assays were conducted with a completely randomized design layout with triplicates per treatment under laboratory conditions. The fungal endophytes used as mycofumigants, Diaporthe sp. and Fusarium proliferatum, were comparatively effective in controlling the anthracnose of banana and mango. The mycelial growth for in vitro assay of Colletotrichum musae and Colletotrichum gloeosporioides were significantly reduced compared to the control when exposed to volatile organic compounds produced by the fungal endophytes. In addition, the results of the in vivo mycofumigation assay against C. musae showed Diaporthe sp. had a significantly higher inhibition rate (93%) than F. proliferatum (67%) when grown on potato dextrose agar plates. The bioefficacy of the Diaporthe sp. and F. proliferatum was improved when grown on peanut and corn seed substrates with a reduction of banana and mango anthracnose severity ranging from 82 to 100%. Cultures grown on corn substrate performed better than those grown on peanut. Pathogenicity tests also revealed that these endophytic fungi did not cause disease in the banana or mango fruits indicating their potential as good biocontrol agents against C. musae and C. gloeosporioides.

Identification and characterization of Colletotrichum species associated with anthracnose disease of banana

AbstractBanana (Musa spp.) is one of the five most abundantly produced fruits in the world and is widely planted in tropical and subtropical areas. Banana anthracnose is one of the main diseases during the growth and postharvest storage period of banana, seriously affecting quality and production. In this study, 24 samples of banana anthracnose were collected near the cities Nanning, Qinzhou, Baise, and Chongzuo in Guangxi Province, China. Based on colony features, conidial and appressorial morphology, and sequence analysis of several genomic regions (internal transcribed spacer [ITS] region, glyceraldehyde‐3‐phosphate dehydrogenase [GAPDH], actin [ACT], β‐tubulin [TUB2], chitin synthase [CHS‐1], calmodulin [CAL], and the intergenic region of apn2 and MAT1‐2‐1 [ApMAT]), the 32 Colletotrichum isolates obtained were identified as five species: C. fructicola (41%), C. cliviicola (28%), C. siamense (16%), C. karstii (9%), and C. musae (6%). A conidial suspension (106 spores/ml) was used to inoculate banana seedlings for pathogenicity tests by applying 20 μl to wound sites. Lesions caused by C. musae developed most rapidly while those of C. karstii took the longest time to develop. This is the first report of C. siamense, and C. karstii associated with banana anthracnose in China, and the first report of C. fructicola and C. cliviicola associated with banana anthracnose worldwide.

First Record of Colletotrichum gloeosporioides Causing Anthracnose of Banana in Pakistan.

Banana (Musa spp.) is one of the most widely grown and consumed fruits in Pakistan and all around the world due to their distinct aroma and taste. In 2018, anthracnose symptoms were observed on banana fruit harvested from different plantations of Sindh- a major banana producing Province of Pakistan. Approximately, 25% of banana fruit collected from different plantations were infected. The symptoms consisted of small brown to reddish-brown spots on the fruit surface and then became sunken lesions as the disease progressed. To identify the pathogen, infected tissues (5 mm in diameter) from the margin of the lesions were surface sterilized by dipping in 1% sodium hypochlorite (NaOCl) for 2 min, 70% ethanol for 30 s, and then rinsed twice with sterile distilled water, plated onto potato dextrose agar (PDA), and incubated at 27°C for 5 days with 12 h light and darkness cycle. Colonies with a similar pattern were consistently isolated and all colonies were sub-cultured using the single-spore method. Colonies first appeared with white colored mycelium and later turned to dark gray. Conidia produced in acervuli were cylindric, hyaline, straight, and aseptate, with both ends rounded. Conidia measured 14.0 ± 0.5 × 3.4 ± 0.6 μm. Conidiomata were dark brown and spherical. On the basis of morphological characterization, the pathogen was identified as Colletotrichum gloeosporioides (Penz.) Penz. & Sacc. (Weir et al. 2012). Two independent isolates (PDL2031 and PDL2032) were used for further genetic analysis. The internal transcribed spacer (ITS) region and chitin synthase 1 (CHS-1) gene were amplified from genomic DNA using primer pairs of ITS1/ITS4 and CHS-79F/CHS-345R, respectively (White et al. 1990; Damm et al. 2012). The GenBank accession numbers (MW493198, MW504711 for ITS and MW530421, MW530422 for CHS-1) of the sequences exhibited 99% to 100% identity to multiple sequences of C. gloeosporioides. To conduct a pathogenicity test, 10 healthy fruits were selected and surface sterilized with 70% ethanol followed by a wash of sterilized water. The fruits were stabbed with a sterile needle and a drop of 20 µl of spore suspension (106 spores/ml) was placed on each wound independently. Meanwhile 10 fruits inoculated with sterile water were treated as controls. The fruits were incubated at 27°C with 90% relative humidity for 10 days. Inoculated fruits exhibited symptoms similar to the original infection. No visible lesions appeared on control fruit. C. gloeosporioides was successfully reisolated from the inoculated fruit, confirming Koch's postulates. Anthracnose of banana is known to be caused by C. musae, C. gloeosporioides, C. siamense, C. tropicale, C. chrysophilum, C. theobromicola, and C. scovillei (Kumar et al. 2017; Peres et al. 2001; Vieira et al. 2017; Zakaria et al. 2009; Zhou et al. 2017). To our knowledge, this is first report of anthracnose of banana caused by C. gloeosporioides in Pakistan. The new disease primarily reduces the quality and yield of Banana. Effective measures should be taken to manage this disease.

First record of Colletotrichum alienum Causing postharvest Anthracnose disease of mango fruit in China.

Mango (Mangifera indica L.) is one of the world's most significant economic fruit crops, and China is the second-largest producer of mango (Kuhn et al., 2017). Postharvest mango anthracnose is caused by Colletotrichum species and reduce the self-life of mature fruit (Wu et al., 2020). Colletotrichum species also cause postharvest anthracnose and fruit rot disease of Apple, Banana and Avocado (Khodadadi et al., 2020; Vieira et al., 2017; Sharma et al., 2017). In July 2019, mango fruits cv. 'Jin-Hwang' were observed at different fruit markets (39°48'42.1"N 116°20'17.0"E) of the Fengtai district, Beijing, China, exhibiting typical symptoms of anthracnose including brown to black lesions in different size (≤ 2 cm) with identified border on the mango fruit surface. Later, the lesions were coalesced and extensively cover the surface area of the fruit. The lesions were also restricted to peel the fruit and pathogen invaded in the fruit pulp. About 30% of mango fruits were affected by anthracnose disease. The margins of lesions from infected mango fruits (n=56) were cut into 2 × 2 mm pieces, surface disinfected with NaClO (2% v/v) for 30 s, rinsed thrice with distilled water for 60s. These pieces were placed on PDA medium and incubated at 25°C for 7 days. Pure culture of fungal isolates was obtained by single spore isolation technique. Initially, the fungal colony was off white, and colony extended with time, turning light gray at the center. The morphological examination revealed that conidia were hyaline, oblong, and unicellular. The conidia were measured from 10 days old culture and dimensions varied from 13.3 to 15.8 µm in length and 4.6 to 6.1 µm in width. For molecular identification, a multi-locus sequence analysis; the Internal Transcribed Spacers (ITS) region, partial actin (ACT) gene, glyceraldehyde-3-phosphate dehydrogenase (GAPDH) gene and chitin synthase (CHS-1) gene were amplified by using the primer sets ITS1/4 (White et al. 1990), ACT-512F/ACT-783R (Carbone and Kohn 1999), GDF1/GDR1 (Guerber et al. 2003) and CHS1-79F/CHS-1-354R (Carbone and Kohn 1999) respectively. The partial sequences of MTY21 were deposited to GenBank accessions (MT921666 (ITS), MT936119 (ACT), MT936120 (GAPDH) and MT936118 (CHS-1). All obtained sequences showed 100% similarity with reported sequences of Colletotrichum alienum ICMP.18691 with accessions numbers JX010217 (ITS), JX009580 (ACT), JX010018 (GAPDH) and JX009754 (CHS-1) which represented the isolate MTY21 identified as C. alienum by constructing Maximum Likelihood phylogenetic tree analysis using Mega X (Kumar et al., 2018). For the confirmation of Koch's postulates, the pathogenicity test was conducted on 36 fresh healthy mango fruits for each treatment. Fruits were punctured with the help of a sterilized needle to create 2mm2 wounds and inoculated with 10µL inoculum (107 spores/mL) of MTY21. Control mango fruits were inoculated with 10µL sterilized distilled water and incubated at 25 °C with 90% relative humidity. The lesions appeared at the point of inoculation and gradually spread on the fruit surface after 7 days post inoculation. The symptoms were similar to the symptoms on original fruit specimens. The re-isolated fungus was identified as C. alienum based on morphological and molecular analysis. Mango anthracnose disease caused by several Colletotrichum species has been reported previously on mango in China (Li et al., 2019). Liu et al. (2020) reported C. alienum as the causal organism of anthracnose disease on Aquilaria sinensis in China. C. alienum has been previously reported causing mango anthracnose disease in Mexico (Tovar-Pedraza et al., 2020) To our knowledge, this is the first report of C. alienum causing postharvest anthracnose of mango in China. The prevalence of C. alienum was 30% on mango fruit which reflects the importance of this pathogen as a potential problem of mango fruit in China.

Antifungal sesquiterpenes with post-harvest anthracnose control effect on bananas from the fungus Fusarium lateritium

To search for antifungal leads, the metabolites of an insect-derived fungus Fusarium lateritium ZMT01 were investigated, providing five sesquiterpenes (1–5), including new molecules microsphaeropsisins D and E (1 and 2). The evaluated antifungal activities in vitro which are higher than the positive control triadimefon include: 1 and 2 towards Fusarium oxysporum (MICs 50, 25 mg L−1; triadimefon 100 mg L−1); 1, 2, 4 and 5 towards Penicillium italicum (MICs 25, 12.5, 25, 25; triadimefon 50 mg L−1), 1, 2 and 4 towards Colletotrichum musae (MICs 25, 12.5, 25; triadimefon 80 mg L−1), 2 and 4 towards Fusarium graminearum (MICs 100, 100; triadimefon 150 mg L−1). The bioassay in vivo displayed that the banana anthracnose control effect of 2 (100 mg L−1) was also higher than that of triadimefon (Inhibition ratios 27.5 ± 2.5%, 55.3 ± 1.4%, 52.1 ± 1.3% for 2, 22.5 ± 2.1%, 47.2 ± 2.0%, 36.6 ± 2.2% for triadimefon at 4 d, 8 d and 12 d, respectively).