Postharvest Fruit Rot Research Articles

Postharvest fruit rot of Bell pepper (Capsicum annuum L.): Pathogenicity and Host range of Alternaria alternata

Bell pepper is a fleshy vegetable cultivated in many countries. A severe fruit rot symptom was observed on green bell pepper in the local vegetable market in New Delhi, India. Fruit rot-associated fungal pathogen isolated on culture medium showed greyish-to-olive green mycelium with prolific conidial production. The pathogenicity test on green bell pepper indicated that inoculated fungus remained quiescent for ten days, triggering typical fruit rot upon the physiological maturation of bell pepper. Based on microscopic features and sequencing of genomic loci such as ITS, Alt a1, and RPB2, the pathogen identity was established as Alternaria alternata. Annotated nucleotides sequence of three genes were deposited and assigned NCBI GenBank accession numbers. Phylogenetic analysis using the taxonomic species of Alternaria strains revealed that the isolate A. alternata Cap_Aa clustered with reference strains of Alternaria alternata. A host range study of A. alternata Cap_Aa confirmed its pathogenicity on tomatoes and eggplant. Results presented in the present work provide knowledge on pathogen identity, host range, and genetic-relationship for developing suitable fruit rot management strategies and postharvest handling in the future.

Pest categorisation of Neoscytalidium dimidiatum.

The EFSA Plant Health Panelperformed a pest categorisation of Neoscytalidium dimidiatum, a clearly defined plant pathogenic fungus of the family Botryosphaeriaceae. The pathogen affects a wide range of woody perennial crops and ornamental plants causing symptoms such as leaf spot, shoot blight, branch dieback, canker, pre- and post-harvest fruit rot, gummosis and root rot. The pathogen is present in Africa, Asia, North and South America, and Oceania. It has also been reported from Greece, Cyprus and Italy, with a restricted distribution. Nevertheless, there is a key uncertainty on the geographical distribution of N. dimidiatum worldwide and in the EU, because in the past, when molecular tools were not available, the two synanamorphs of the pathogen (Fusicoccum-like and Scytalidium-like) might have been misidentified based only on morphology and pathogenicity tests. N. dimidiatum is not included in Commission Implementing Regulation (EU) 2019/2072. Because of the wide host range of the pathogen, this pest categorisation focuses on those hosts for which there is robust evidence that the pathogen was formally identified by a combination of morphology, pathogenicity and multilocus sequence analysis. Plants for planting, fresh fruits and bark and wood of host plants as well as soil and other plant growing media are the main pathways for the further entry of the pathogen into the EU. Host availability and climate suitability factors occurring in parts of the EU are favourable for the further establishment of the pathogen. In the areas of its present distribution, including Italy, the pathogen has a direct impact on cultivated hosts. Phytosanitary measures are available to prevent the further introduction and spread of the pathogen into the EU. N. dimidiatum satisfies the criteria that are within the remit of EFSA to assess for this species to be regarded as potential Union quarantine pest.

Postharvest fruit rot of pear caused by Diaporthe eres in China

Postharvest fruit rot of pear caused by Diaporthe eres in China

First Report of Penicillium oxalicum Causing Postharvest Fruit Rot on Citrus in China.

Ougan (Citrus reticulata cv. Suavissima) is a distinct citrus cultivar local to Zhejiang province, China. (Guo et al. 2021). In November 2021, an unknown postharvest fruit rot was observed in the Sanyang wetland Ougan planting area, Wenzhou City of Zhejiang Province (27.96 °N, 120.69 °E). About 3% of diseased fruits with similar fruit rot symptoms were observed in 900 mandarin fruits from four commercial storages. Initially, the symptoms appeared as light brown lesions that turned deep brown as the lesions expanded. To identify the pathogen, segments (5 mm2) from margins of rotted tissue were excised from 5 symptomatic fruits, surface disinfested twice with 75% ethanol, rinsed three times with sterilized water, placed aseptically onto potato dextrose agar (PDA) medium and incubated for 7 days at 25℃ in darkness for identification. Five fungal isolates with the same morphology were obtained using the single spore method (Leslie and Summerell 2006). Initially, the fungus produced fluffy and white aerial mycelium that eventually turned green on PDA medium after 3 days. Conidiophores were broom-shaped (17.5 ± 2.5 μm) (n=50). Conidia were unicellular and ellipsoid (3.5 to 5.0 ×2.5 to 4.0 μm) (n=50). These morphological characteristics were consistent with Penicillium species (Wu et al. 2022). WZU-OG1 was chosen as a representative isolate for further study. For molecular identification, PCR amplification and DNA sequencing were performed using primers ITS1/ITS4 (White et al. 1990), bt2a/bt2b (Glass and Donaldson 1995), and RPB2-5F/RPB2-7R (Liu et al. 1999) to amplify the complete internal transcribed spacer (ITS) region, β-tubulin gene (TUB), and a portion of RNA polymerase second largest subunit (RPB2). The ITS, TUB, and RPB2 gene sequences of isolate WZU-OG1 were deposited in the GenBank database with acc. nos. ON332735, ON428245, and ON524171, respectively. BLASTn analysis respectively showed 561/561 (MH855125), 414/427 (KF296462), and 936/954 (JN121456) matching with Penicillium oxalicum CBS 219.30. A neighbor-joining phylogenetic analysis based on the concatenated nucleotide sequences (ITS, TUB, and RPB2) grouped this isolate in the Penicillium oxalicum species complex clade at 100% bootstrap support. To verify pathogenicity, 20 healthy mandarin fruit of cultivar Sanyang were superficially disinfested with 75% ethanol and then washed with distilled water. A conidial suspension of 1 × 105 conidia/ml from a 5-day-old culture of WZU-OG1 was injected into 10 fruits (10 μL per fruit). An equal number of fruits inoculated with sterile water were used as the negative control. The inoculated fruits were stored in a constant temperature incubator under the conditions of 28 ℃, 90% humidity, and incubated in a 12-h light/12-h dark cycle for 12 days. Symptoms similar to those on the naturally infected fruit began 5 days after inoculation, whereas no symptoms occurred on the controls. The experiment was repeated three times, and similar symptoms were observed in all diseased fruits. Then, the fungus was reisolated from these infected fruits and identified as P. oxalicum by the morphological and molecular methods described above. This is the first report of P. oxalicum causing postharvest mandarin decay and this study will enable us to rapidly diagnose this disease, identify the occurrence of this disease and develop adequate management strategies to control it in China.

The Blueberry Fruit Mycobiome Varies by Tissue Type and Fungicide Treatment

The microbial ecology of agricultural products may provide crucial insights into the management of postharvest fruit rots. To investigate postharvest microbial communities of highbush blueberry ( Vaccinium corymbosum), five fungicide spray programs were evaluated for their influence on the mycobiome of fruit skin and within the fruit pulp. The mycobiome was characterized by sequencing amplicons of the internal transcribed spacer 1 (ITS1) region with primers ITS1f and ITS4 with the Illumina MiSeq 300bp v3 system. Two of the five programs utilized commercial biological fungicides, two utilized azoxystrobin, and one utilized a series of treatments to simulate a realistic disease management program. Fungicide applications reduced diversity of the fruit skin mycobiome ( R2 = 0.409, P = 0.0001) and had a moderate impact on the pulp mycobiome ( R2 = 0.233, P = 0.0001). The mycobiome of the fruit pulp was also more variable than the skin mycobiome. In comparison with the untreated controls, each fungicide treatment program had a strongly significant effect on the β diversity of the blueberry fruit skin mycobiome ( R2 = 0.53 to 0.73, P = 0.0001). In the pulp, three of the five treatments had moderate but significant effects on β diversity in comparison with the control ( R2 = 0.10 to 0.18, P = 0.0005 to 0.017). Most samples indicated that fungi belonging to genera Epicoccum, Papiliotrema, and Sporobolomyces were widely prevalent and abundant across treatments and tissues. Fruit pathogen Botrytis cinerea was particularly abundant in the pulp of three of the fungicide treatments. Results from this study provide a baseline for future exploration of postharvest rot pathology and provide a community context on how fungicides may alter fungal communities in agricultural systems.

Fungal pathogens causing postharvest fruit rot of wolfberry and inhibitory effect of 2,3-butanedione.

Fungal pathogen contamination is one of the most important factors affecting the postharvest quality and shelf life of wolfberry fruits. Therefore, the prevention and control of fungal pathogens that cause fruit rot has become particularly important. Volatile antifungal agents of biological origin have broad application prospects. They may be safer and more efficient than traditional physical and chemical methods. Four pathogenic fungi were isolated and purified from rotting wolfberry. These pathogenic fungi were determined to be Mucor circinelloides LB1, Fusarium arcuatisporum LB5, Alternaria iridiaustralis LB7, and Colletotrichum fioriniae LB8. In vitro fumigation experiments showed that 2,3-butanedione can effectively inhibit the mycelial growth, spore germination, and sporulation ability of pathogenic fungi. The scanning electron microscope (SEM) showed morphological changes in hyphae. Propidium iodide (PI) Staining and leakage of 260 and 280 nm-absorbing increased, suggesting damage to cell membranes. Furthermore, 2,3-butanedione was found to significantly improve fruit firmness, soluble solid, total phenol, flavonoid, and soluble sugar content, as well as higher SOD enzyme activity and lower PPO and POD enzyme activity in the treated fruit, indicating that 2,3-butanedione can effectively reduce the adverse effects of pathogenic fungi in wolfberry. Based on these results, we conclude that 2,3-butanedione is effective against infection by pathogenic fungi in post-harvest wolfberry. 2,3-butanedione should be considered a viable substitute for conventional fungicides that are currently used to control rot in wolfberry.

Pest categorisation of Lasiodiplodia pseudotheobromae.

The EFSA Plant Health Panelperformed a pest categorisation of Lasiodiplodia pseudotheobromae, a clearly defined fungus of the family Botryosphaeriaceae, which was first described in 2008 as a cryptic species within the L. theobromae complex. The pathogen affects a wide range of woody perennial crops and ornamental plants causing root rot, damping-off, leaf spots, twig blight, cankers, stem-end rot, gummosis, branch dieback and pre- and post-harvest fruit rots. Lasiodiplodia pseudotheobromae is present in Africa, Asia, North and South America and Oceania and has also been reported from Spain with a restricted distribution. However, there is uncertainty on the status of the pathogen worldwide and in the EU because in the past, when molecular tools (particularly multigene phylogenetic analysis) were not available, the pathogen might have been misidentified as L. theobromae. Lasiodiplodia pseudotheobromae is not included in Commission Implementing Regulation (EU) 2019/2072 and there are no interceptions in the EU. Because of the very wide host range of the pathogen, this pest categorisation focused on those hosts for which there is robust evidence that the pathogen was formally identified by a combination of morphology, pathogenicity and multilocus sequence analysis. Plants for planting, including seeds, fresh fruits and bark and wood of host plants as well as soil and other plant-growing media are the main pathways for the further entry of the pathogen into the EU. Host availability and climate suitability factors occurring in parts of the EU are favourable for the further establishment of the pathogen. In the area of its present distribution, including Spain, the pathogen has a direct impact on cultivated hosts. multilocus measures are available to prevent the further introduction and spread of the pathogen into the EU. Lasiodiplodia pseudotheobromae satisfies the criteria that are within the remit of EFSA to assess for this species to be regarded as potential Union quarantine pest.

First Report of Neopestalotiopsis clavispora Causing Postharvest Fruit Rot on Actinidia arguta in Liaoning Province, China.

Actinidia arguta, commonly called hardy kiwifruit or kiwiberry, is a perennial vine of Actinidiaceae Actinidia genus. Understanding the main pathogens that cause the fruit rot of A. arguta during storage is of great significance for finding strategies to prevent fruit rot. In September 2020, the A. arguta (Sieb.et Zucc.) Planch. ex Miq. LD133 was harvested from a farm in Dandong City, Liaoning Province, China (40°31'N, 124°20'E). After being stored at room temperature for about a week, the fruit rotted (no mechanical damage or wound). Initial symptoms were localized irregular spots, which then became soft and the spots connected into large, flaky, light brown lesions. Later symptoms were dark brown lesions and rot that affect the entire fruit. The strain that shows the same morphology as observed in the four decaying tissues was isolated on PDA. The strain was white, edge irregular and surface wavy, and the reverse side was pale yellow (Fig. S1A). The black viscous acervuli appeared on the surface of the mycelium after 7 to 10 days at 25℃ with 12 hours photoperiod. Conidia were fusiform to ellipsoid, straight to slightly curved, 5.7×25.9 µm (width × length), n=30, with five versicolor cells (three brown median cells, two hyaline cells on apical and basal). The apical cell generally contains one to four appendages on conidia (Fig. S1B). For identify the selected strains, three genetic regions (ITS, TUB and TEF 1-α) were used for amplification and sequencing. These sequences of pathogen shared 98 to 100% homology with Neopestalotiopsis clavispora. Then, a phylogenetic tree was constructed by the Bayesian algorithm using PhyloSuite (v1.2.2) (Zhang et al. 2020). Based on the morphological and molecular characterization, the pathogen was identified as Neopestalotiopsis clavispora (Chamorro et al. 2016). Next, pathogenicity of the screened strains was determined by wound inoculation method. Ripe healthy fruits are immersed in 1% NaClO, rinsed twice with sterile water. A sterile needle was used to penetrate 1-2mm of peel, and then inoculate hyphae (about 5mm in length, 1mm in diameter), and 10 µL of sterile water as a control. The treated fruits are stored in an artificial climate chamber (22°C, 70% relative humidity with 12 hours photoperiod). The inoculated fruit began to show signs of rot on the second day, and after 6 days, lesions similar to those found during storage of the pathogenic isolated fruit appeared. Similarly, 10 µL (106 conidia/mL) conidial suspension exhibited decay symptoms. Compared with conidial suspension as inoculum, hyphae has shorter incubation period and stronger pathogenicity to fruit (Fig. S2). The pathogen was re-isolated from these infected fruits and identified as N. clavispora, thus fulfilling Koch's postulates. N. clavispora has been reported causing root and crown rot on strawberry in Spain, Argentina, Uruguay and Italy (Chamorro et al. 2016; Gilardi et al. 2019; Machin et al. 2019; Obregon et al. 2018), and as a pathogen on blueberry in Spain and Korea (Borrero et al. 2018; Lee et al. 2019), and causing leaf spot on macadamia in Brazil (Santos et al. 2019), and causing leaf spot on Syzygium cumini in India (Banerjee and Rana 2020). However, to our knowledge, this is the first report of Neopestalotiopsis clavispora causing postharvest fruit rot of A. argute in the worldwide. The identification of the pathogen is of great significance for conducting research on A. argute fruit preservation to prolong its shelf life and improve its merchantability.

Mango germplasm screening for the identification of sources of tolerance to anthracnose

Colletotrichum species are one of the most common causes of postharvest fruit rot in mango in Australia, particularly in the tropical region of north Queensland, and can result in significant losses if not managed. The research aims were to identify sources of anthracnose tolerance and to determine if host material other than fruit could improve or fast track the screening process and result in improved breeding efficiency. Access to the Australian National Mango Genebank (ANMG) collection enabled fruit screening of more than 100 Mangifera indica cultivars or Mangifera species for tolerance to anthracnose by artificial inoculation with Colletotrichum asianum over a period of 14 years. Mean lesion diameters were compared with those on a known susceptible M. indica cultivar Kensington Pride (KP) and a tolerant M. laurina cultivar Lombok. Inoculation of leaf discs and entire leaves was evaluated in the laboratory and the field as alternative assays for tolerance to anthracnose and was assessed by presence/absence of disease. Screening of fruit has shown that anthracnose tolerance within the mango germplasm is highly variable and needs to be assessed over multiple years. None of the alternative laboratory bioassays provided consistent or reliable data. The in-field artificial inoculation of immature leaf flush was successful but was not deemed suitable for adoption due to practical restraints. While resistance to anthracnose in fruit has not yet been identified, some cultivars and Mangifera spp. showed promise for inclusion as parents in future breeding programs.

Impact of Ozonated Water on Brown Rot Development and Storage Potential of Nectarine and Plum

ABSTRACT Postharvest fruit rot is one of the most serious diseases of fleshy stone fruits such as nectarine and plum. The efficacy of ozonated water sprayed in different doses in a packing line [low dose, 1.5 ppm ozone (Ο3) for 100 sec; medium dose, 3.0 ppm Ο3 for 100 sec; high dose, 1.5 ppm Ο3 for 400 sec] in controlling the brown rot (Monilinia fructicola) development was studied in the nectarine cultivar ‘Τasty Free’ and the plum cultivar ‘Angelino.’ Changes in flesh firmness, ripening index, ethylene and respiration rate, and peel color during storage (0℃ for 3 weeks) and shelf-life (20℃ for 4 days) were used as ‘indicator’ quality parameter responses in storage and shelf-life. The most efficient dose of O3 in controlling the Monilinia infection was the highest applied, followed by the middle O3 dose. The efficacy of O3 treatments was higher when a lower Monilinia spore concentration was applied. The O3 doses applied did not cause any visible symptoms of O3 toxicity, yet they caused an acceleration in the fruit ripening process, suggested mainly by an increase in the fruit respiration rate in both nectarine and plum, and an increase in the ripening index and darker peel coloration only in plums. In conclusion, spraying with ozonated water is an eco-friendly and efficient sanitizing agent, but in fleshy commodities such as nectarine and plum may diminish their postharvest performance.

Early warning of Diaporthe infection in kiwifruit soft rot by plasmonic dimer-enhanced Raman spectroscopy.

Diaporthe genus is the dominant pathogens of kiwifruit soft rot with long incubation period and rapid onset and very hard to detect in advance. It is of great significance to develop point-of-care tests for disease prevention and field management. Here we screen a pair of genus-level primers for constructing plasmonic dimer Rayleigh/Raman spectroscopy, which enables rapid, specific, and sensitive detection of Diaporthe genus in real kiwifruits. Dark-filed Rayleigh scattering clearly visualizes the target-induced dimer assembly of nanoprobes. Plasmonic dimer-enhanced Raman spectroscopy has high specificity for 12 common isolates of Diaporthe genus in one batch samples with highly accurate results and much higher sensitivity than polymerase chain reaction (PCR). It realizes the recognition of Diaporthe infection at an early stage of 24h when the kiwifruits do not have any noticeable symptoms. It demonstrates a bright prospect of point-of-care tests for early warning of kiwifruit soft rot infection and quality control.

First Report of Trichothecium roseum Causing Postharvest Fruit Rot on Purple Passion Fruit in China.

Purple Passion fruit (Passiflora edulis) is widely cultivated in many regions of southern China as an edible tropical fruit with excellent nutritional value and high economic value. In July 2021, postharvest fruit rot was observed on 20-25% of purple passion fruit in several fruit markets of Dehong City in Yunnan Province. Symptoms on infected fruits were irregular, pink-brown, soft, and water-soaked lesions, which enlarged and formed sunken patches with time as well as producing a small amount of white mycelium. To isolate the causal organism, five diseased fruits were collected from different fruit markets. A conidial mass from an individual sorus observed on an infected fruit was isolated and cultured on potato dextrose agar (PDA) supplemented with 50 μg ml-1 of streptomycin, and five fungal isolates were obtained. These isolates were morphologically similar and produced pale pink colonies on PDA for 7 days containing several conidiophores with abundant conidia. Mycelia were hyaline, 2 µm in diameter, and conidiophores were simple or branched (100 to 286 × 1.5 to 2.5 µm, n=50). Conidia were pyriform, ovate, with papillary protuberances at one end. Almost all conidia were two-celled and single-septate (5.8 to 9.1 × 1.7 to 4.9 µm, n=50). The morphology of the fungi resembled Trichothecium roseum as reported previously (Inácio et al. 2011). To further confirm the fungal species, isolate PASF4 was selected for molecular identification by amplifying and sequencing the ribosomal internal transcribed spacer (ITS) and large subunit (LSU) genes. Primers and PCR amplification were described by Fell et al. (2000). Results showed that both the ITS (GenBank accession OL336243) and LSU (OL336242) gene sequences had 100% similarity to T. roseum in NCBI database (MH856757 and MH868278). Maximum likelihood tree was constructed using MEGA 7 (Felsenstein, 1981) based on concatenated sequences (ITS and LSU) of isolate PASF4 and reference strains. Phylogenetic analysis showed that isolate PASF4 belonged to T. roseum clade. Based on morphological characteristics and phylogenetic analysis, isolate PASF4 was identified as T. roseum (Inácio et al. 2011). To confirm their pathogenicity, healthy purple passion fruits (cv. Tainong-1) were disinfected in 0.5% NaClO solution for 2 min, and then washed with sterile water. After wounding with a sterile needle, the fruits were inoculated by placing mycelium agar plugs on the wounds, and mock inoculation with mycelium-free PDA plugs served as control. Five fruits were used in each treatment. All fruits were maintained in plastic boxes at 25 °C. Disease symptoms appeared after inoculation for 4-7 days on all inoculated fruits, which were similar to those observed in fruit markets. No symptoms were observed on fruits used as control. The Trichothecium isolates were re-isolated from symptomatic fruits thus fulfilling Koch's postulates. Trichothecium roseum has been reported to cause fruit rot of tomato, apple and orange in Pakistan (Hamid et al., 2014) and fruit rot of pepper in China (Lin et al., 2016). To our knowledge, this is the first report of T. roseum causing fruit rot on purple passion fruit worldwide, and these data will provide useful information for developing effective control strategies.

First report of Fusarium ipomoeae and F. citri causing postharvest fruit rot of winter squash (Cucurbita maxima)

First report of Fusarium ipomoeae and F. citri causing postharvest fruit rot of winter squash (Cucurbita maxima)

Morphology, Molecular Identification, and Pathogenicity of Two Novel Fusarium Species Associated with Postharvest Fruit Rot of Cucurbits in Northern Thailand.

Fruit rot of cucurbits caused by several pathogenic fungi has become an important postharvest disease worldwide. In 2022, fruit rot on watermelon (Citrullus lanatus) and muskmelon (Cucumis melo) was observed during the postharvest storage phase in the Chiang Mai and Phitsanulok Provinces of northern Thailand. These diseases can lead to significant economic losses. This present study was conducted to isolate the causal agent of fungi in lesions of fruit rot. A total of four fungal isolates were obtained, of which two isolates (SDBR-CMU422 and SDBR-CMU423) were obtained from rot lesions of watermelons, while the remaining isolates (SDBR-CMU424 and SDBR-CMU425) were obtained from rot lesions of muskmelons. All fungal isolates were identified using both morphological characteristics and molecular analyses. Morphologically, all isolated fungal isolates were classified into the genus Fusarium. Multi-gene phylogenetic analyses of a combination of the translation elongation factor 1-alpha (tef-1), calmodulin (cam), and RNA polymerase second largest subunit (rpb2) genes reveled that four fungal isolates belonged to the Fusarium incarnatum-equiseti species complex and were distinct from all other known species. Thus, we have described them as two new species, namely F. citrullicola (SDBR-CMU422 and SDBR-CMU423) and F. melonis (SDBR-CMU424 and SDBR-CMU425). A full description, illustrations, and a phylogenetic tree indicating the position of both new species have been provided. Moreover, pathogenicity tests were subsequently performed and the results showed that F. citrullicola and F. melonis caused symptoms of fruit rot on inoculated watermelon and muskmelon fruits, respectively. Notably, this outcome was indicative of the symptoms that appeared during the postharvest storage phase. To our knowledge, two new pathogenic fungi, F. citrullicola and F. melonis, are new causal agents of watermelon and muskmelon fruit rot, respectively. Importantly, these findings provide valuable information for the development of effective strategies for the monitoring and prevention of these diseases.

Fusarium falciforme and F. oxysporum causing postharvest fruit rot of watermelon (Citrullus lanatus) in Malaysia: A first report

Watermelon is one of the fruit crops being produced in large scale (9000 ha in the year of 2021) in Malaysia. However, fungal diseases are a major factor in suppressing yield. In November 2019, field surveys in the states of Kelantan, Terengganu, and Penang found harvested watermelon showing fruit rot disease symptoms. In the present study, we delineated the pathogens causing these symptoms using morphological observations, molecular methods, and pathogenicity tests. Morphological characterization, DNA sequencing and phylogenetic analysis using ITS and TEF loci revealed that two isolates, S9 and S11, isolated from symptomatic fruits were Fusarium falciforme and F. oxysporum, respectively. Pathogenicity tests confirmed the both Fusarium species caused the observed symptoms, fulfilling Koch's postulates. This is the first report of these Fusarium species as the pathogens of watermelon fruit rot in Malaysia. Findings from the present study contribute to the knowledge of watermelon diseases in Malaysia, provide an essential foundation for development of effective pre-harvest and postharvest disease management strategies to curb disease spread.

‘Corrigendum to “Development of eco-friendly nanoemulsions of some natural oils and evaluating of its efficiency against postharvest fruit rot fungi of cucumber” [Ind. Crops Prod. 159 (2021) 113049

‘Corrigendum to “Development of eco-friendly nanoemulsions of some natural oils and evaluating of its efficiency against postharvest fruit rot fungi of cucumber” [Ind. Crops Prod. 159 (2021) 113049

Profiling the secretomes of Penicillium expansum reveals that a serine carboxypeptidase (PeSCP) is required for the fungal virulence on apple fruit

Phytopathogenic fungi often secrete effector proteins that suppress host immunity and facilitate infection. Penicillium expansum is an important postharvest fruit rot pathogen that caused blue mold. However, little is known about the function of effectors secreted by P. expansum. In this study, a LC-MS/MS based proteomic approach was used to analyze the secretomes of P. expansum grown in apple juice, a medium used to mimic natural in planta condition. A total of 103 and 148 effector candidates were identified after incubation for 24 and 72 h, respectively. As expected, the majority of the proteins identified with known function were hydrolytic enzymes, such as cell-wall degrading enzymes and peptidases/proteases. One of the identified effector candidates is a serine carboxypeptidase (PeSCP) which was detected at 24 h after incubation and significantly up-regulated at the early stage of infection on apple fruit. Deletion of PeSCP gene led to reduced conidiation, germination as well as affected fungal growth and morphology, and also resulted in altered tolerance to environmental stresses, greatly reduced extracellular carboxypeptidase activity and virulence of P. expansum on apple fruit. Scanning electron microscopy revealed that the knockout of PeSCP in P. expansum impaired its ability to penetrate host plant tissues. Together, our findings demonstrate an important role of PeSCP in the virulence of P. expansum on apple fruit.

Ethyl acetate produced by Hanseniaspora uvarum is a potential biocontrol agent against tomato fruit rot caused by Phytophthora nicotianae.

In this study, an oomycete strain FQ01 of Phytophthora nicotianae, which could cause destructive postharvest disease, was isolated. At present, chemical fungicides are the main reagents used for controlling Phytophthora diseases. It is necessary to find new control techniques that are environmentally friendly. The biocontrol activity of Hanseniaspora uvarum MP1861 against P. nicotianae FQ01 was therefore investigated. Our results revealed that the volatile organic compounds (VOCs) released by the yeast strain MP1861 could inhibit the development of P. nicotianae FQ01. The major component of the VOCs produced by the yeast strain MP1861 was identified to be ethyl acetate (70.8%). Biocontrol experiments showed that Phytophthora disease in tomato fruit could be reduced by 95.8% after the yeast VOCs treatment. Furthermore, ethyl acetate inhibited the mycelial growth of the oomycete strain FQ01, and damaged the pathogen cell membrane. This paper describes the pioneering utilization of the yeast strain MP1861 for biocontrol of postharvest fruit rot in tomato caused by P. nicotianae.

First Report of Postharvest Fruit Rot in Solanum muricatum Aiton Caused by Alternaria alternata in southwest China.

Solanum muricatum is native to South America and well known for its sweet, attractive, nutritious fruits. S. muricatum has been cultivated in China since the 1980s and increasingly popular (Li et al. 2015). In November 2021, an unknown fruit rot was observed in Shilin County of Yunnan Province (24.77 °N, 103.28 °E). The incidence of this disease was about 16% of 500 postharvest S. muricatum fruits after 7 d in storage room (25°C, 90% relative humidity). The initial symptoms were small brown spots on the fruit surface, which gradually expanded into irregular brown or black lesions, and gray-white mold developed in the center of the lesions, eventually the fruit turned rot. To isolate the pathogen, ten fruits with typical symptoms were collected and surface-sterilized with 75% ethanol for 45 s. Small fragments (5 × 5 mm) from the margin of lesions on fruit were disinfected with 1% sodium hypochlorite for 60 s, washed three times with sterile water then transferred to potato dextrose agar (PDA), and incubated at 28 ± 1℃ for 3 days (Li et al. 2022). Two fungal isolates with the same morphology were obtained and purified by single-spore isolation method. The colony was covered with thick fluffy aerial mycelia and the center was dark brown or black with white margins. Conidia were brown, pyriform or ellipsoid, with 1 to 3 longitudinal and 2 to 6 transverse septa, 15.12 to 34.01 × 6.90 to 12.73 μm (21.22 × 9.69 μm on average, n=50) in size. These morphological characteristics were consistent with Alternaria alternata (Li et al. 2015; Xiang et al. 2021; Alberto. 1992). For molecular identification, genomic DNA was extracted from a representative isolate, and primers ITS1/ITS4 (Gardes et al. 1993), TEF-F/TEF-R (Lawrence et al. 2013), Alt-F/Alt-R (Hong et al. 2005), GPD-F/GPD-R (Berbee et al. 1999) and EPG-F/EPG-R (Peever et al. 2004) were used to amplify the internal transcribed spacer (ITS), translation elongation factor 1-alpha (TEF), Alternaria major allergen (Alt a1), glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and endo-polygalacturonase (endoPG), respectively. The obtained DNA sequences (ITS: OM049821; TEF: OM069656; Alt a1: OM069655; GAPDH: OM069654 and endoPG: OM069653) showed over 99% homology with that of A. alternata (GenBank Accession No. MN856355.1 (565/573 bp); MN258023.1 (267/267 bp); KY923227.1 (491/501 bp); LC131645.1 (608/609 bp) and MN698284.1 (452/454 bp)). A phylogenetic tree based on the combined ITS, TEF, Alt a1, GAPDH, and endoPG sequences using the maximum likelihood methods with Kimura 2-parameter model, bootstrap nodal support for 1000 replicates in MEGA7.0 (Li et al. 2019) revealed that the isolate was assigned to A. alternata. To confirm pathogenicity, 10 μL spore suspension (1.0 × 106 conidia/ml) obtained from 7-day-old PDA cultures of each isolate were inoculated on 15 needle-wounded and 15 non-wounded surface-disinfected fruits, respectively. Healthy fruits were inoculated with sterile water as controls and the experiment was repeated 3 times. All fruit were incubated at 25 ± 1℃, 90% relative humidity. After 7 days, all the wounded and non-wounded fruit inoculated with A. alternata showed similar symptoms to those observed on the previously fruits, while the control fruits remained healthy. The same pathogen was again isolated from the inoculated fruits, thus Koch's postulates were fulfilled. A. alternata causing fruit rot of Prunus avium and Mangifera indica in China were reported in previous studies (Ahmad et al. 2020; Liu et al. 2019). As far as we know, this is the first report of postharvest fruit rot on S. muricatum caused by A. alternata in southwest China. This work provides a basis for the development of control strategies of the disease in the future.

First Report of Postharvest Fruit Rot Disease of Yellow Peach Caused by Diaporthe eres in China.

The yellow peach (Amygdalus persica), is a fruit crop native to China with golden peel and pulp that is of particular interest in the fruit markets. In August of 2021, yellow peaches showing fruit rot symptoms were purchased from a commercial market in Linyi city, Shandong province, China. The symptoms included circular, tan to brown in color, rotten, necrotic lesions, and whitish mycelium mass in the center of the lesions. The infected fruit were surface disinfected with 1% NaClO for 30 s and rinsed with sterile distilled water three times. Diseased tissues from the infected fruits were cut into small segments, aseptically shifted onto potato dextrose agar media containing petri plates and incubated at 25℃ for 5 days. Eight isolates were obtained in total from two isolation experiments. Fungal colonies were initially white, aerial, fluffy at first, and gradually turned brown to gray, with black stromata at maturity. Alpha conidia were aseptate，hyaline，fusiform to ellipsoidal，and ranged in size from 4.16 to 7.76 µm × 1.95 to 3.14 µm (n=30). Beta conidia were aseptate, hyaline, filiform, curved to hamate, and 15.91 to 22.55 µm × 0.82 to 1.66 µm (n=30). The morphological characteristics were consistent with those of Diaporthe species (Gomes et al. 2013). For further identification, a multigene phylogenetic analysis was carried out. The internal transcribed spacer (ITS) region, translation elongation factor 1-α (TEF1-α), histone H3 (HIS), calmodulin (CAL), and β-tubulin (TUB) genes of two representative isolates were amplified by using primers ITS1/ITS4, EF1-728F/EF1-986R, CYLH3F/H3-1b，CAL228F/CAL737R, and Bt2a/Bt2b (Chaisiri et al. 2021), respectively. The sequences were deposited in GenBank (Accession No. OL375154 for ITS; OL406409 for TEF1-α; OL406410 for HIS; OL106407 for CAL; OL406408 for TUB). phylogenetic analyses were conducted using the concatenation of multiple sequences (ITS, TEF1-α, HIS, CAL, TUB) with Maximum Likelihood (ML) in IQtree v1.5.6 (Nguyen et al. 2015). Based on the morphological and phylogenetic characters, the isolates were identified as D. eres. A Pathogenicity test was performed by wound inoculation on harvested fruits of A. persica Variety 'Jinxiu'. Mature and healthy yellow peaches purchased from Shandong, Anhui, and Hunan Provinces in China were surface sterilized with 1% NaClO solution for 1 minute, rinsed with sterile water and dried. Each fruit was wounded with a sterile scalpel creating a 2-3 mm incision on the peel. A 5 mm agar disc with mycelium grown on PDA at 28℃ for 7 days was placed on wound and sealed with parafilm. Sterile PDA plugs were used as controls. Ten fruit were used for each treatment and the assays were repeated three times. Inoculated fruit were placed in sterilized transparent plastic cans containing wet, sterile paper towels. After 5 days of incubation at 25℃, the same rot symptoms were observed on fruits inoculated with mycelium and the control remained symptomless. D. eres was re-isolated from the lesions of inoculated fruits and the pathogen identification was confirmed by molecular analysis, thus fulfilling complete Koch's postulates. Although D. eres was previously reported on peach trees of causing shoot blight (Thomidis and Michailides 2009) and stem canker (Prencipe et al. 2017). To our knowledge, this is the first report of D. eres causing postharvest fruit rot of yellow peach in China and it may lead to considerable economic losses in the peach industry should post-harvest disease management practices not be implemented.