Neoscytalidium Dimidiatum Research Articles

Overcoming Encouragement of Dragon Fruit Plant (Hylocereus undatus) against Stem Brown Spot Disease Caused by Neoscytalidium dimidiatum Using Bacillus subtilis Combined with Sodium Bicarbonate.

The use of the supernatant from a Bacillus subtilis culture mixed with sodium bicarbonate was explored as a means of controlling stem brown spot disease in dragon fruit plants. In in vitro experiments, the B. subtilis supernatant used with sodium bicarbonate showed a strong inhibition effect on the growth of the fungus, Neoscytalidium dimidiatum, the agent causing stem brown spot disease and was notably effective in preventing fungal invasion of dragon fruit plant. This combination not only directly suppressed the growth of N. dimidiatum, but also indirectly affected the development of the disease by eliciting the dragon-fruit plant’s defense response. Substantial levels of the pathogenesis-related proteins, chitinase and glucanase, and the phenylpropanoid biosynthetic pathway enzymes, peroxidase and phenyl alanine ammonia-lyase, were triggered. Significant lignin deposition was also detected in treated cladodes of injured dragon fruit plants in in vivo experiments. In summary, B. subtilis supernatant combined with sodium bicarbonate protected dragon fruit plant loss through stem brown spot disease during plant development in the field through pathogenic fungal inhibition and the induction of defense response mechanisms.

Deterioration of wood exposed to mangrove ecosystem

Deterioration of three types of wood, namely meranti wood (Shorea spp.), mahoni wood (Swietenia mahagoni) and damar wood (Agathis dammara) in underwater exposure in the mangrove ecosystem investigated. The wood soaked for six months, at a depth of 1.5 meters and salinity 7.3. After six months identification of organisms that attack wood is carried out. The types of organisms found attacking wood samples are Teredo navalis and Balanus sp. The types of fungi found include Paecilomyces victoriae, Scytalidium lignicola, Aspergillus sp., Penicillium corylophilum, Aspergillus fumigatus, Biporalis australiensis, Absidia sp, Aspergillus niger and Phytophthora cryptogea.

Biological prevention and control of pitaya fruit canker disease using endophytic fungi isolated from papaya.

Pitaya fruit canker is an important disease in pitaya production. Facilitating resistance through the application of biological control principles is a promising alternative to traditional control strategies. This study evaluated the induced resistance of Penicillium rolfsii, numbered Y17 isolated from papaya leaves in pitaya fruit, and evaluated the activity of the defense enzymes, total antioxidant capacity (T-AOC), and malondialdehyde (MDA) content of the treated fruit. The results demonstrate that treatment with Y17 effectively induced resistance of pitaya fruit to canker disease caused by Neoscytalidium dimidiatum, with an inhibition rate of 70.87%. In addition, Y17 notably improved the activities of peroxidase, catalase, and polyphenol oxidase as well as the T-AOC of the treated samples. Y17 treatment reduced the MDA content in these fruits. Taken together, our results suggest that Y17 treatment could trigger pitaya fruit defense responses and effectively induce resistance to fruit canker disease.

Identification and pathogenicity of Botryosphaeriaceae species associated with root and stem rot of sweet potato in Brazil

AbstractThe sweet potato (Ipomoea batatas) is characterized by the production of tuberous roots rich in starch and is one of the most produced and consumed vegetables in Brazil. Botryosphaeriaceae, among other fungi, are known to cause root and stem rot of sweet potato. However, no representative and accurate study has been performed for the correct identification of these fungal species in sweet potato in Brazil. Therefore, this study aimed to identify the Botryosphaeriaceae species associated with root and stem rot of sweet potato and confirm their pathogenicity. Tuberous roots and stems of sweet potato with rot symptoms were collected in production fields and markets and used for fungal isolations. The identification of fungi was based on the morphology of reproductive structures and phylogenetic analyses of the gene regions ITS, tef1‐α, and rpb2. The following species were identified: Lasiodiplodia theobromae, L. hormozganensis, Macrophomina phaseolina, M. euphorbiicola, M. pseudophaseolina, and Neoscytalidium dimidiatum. For the pathogenicity test, one representative isolate for each species was inoculated in healthy tuberous roots and in 30‐day‐old healthy seedlings. Black and necrotic lesions on tuberous roots and stems were observed in all replications and resulted in the death of some seedlings. This is the first report of L. hormozganensis, M. pseudophaseolina, and M. euphorbiicola, as causal agents of the stem and root rot of sweet potato and N. dimidiatum as a causal agent of stem rot worldwide.

Comparative proteomic analysis of the fungal pathogen Neoscytalidium dimidiatum infection in the pitaya

Comparative proteomic analysis of the fungal pathogen Neoscytalidium dimidiatum infection in the pitaya

In Vitro Efficiency of Some Fungicides Against Neoscytalı̇dı̇um dı̇mı̇dı̇atum (Penz.) Crous and Slippers Causing Sudden Shoot Dry on Apricot Trees

Turkey is known as a top producer and exporter of apricots, both fresh and dried. More than half of Turkey's apricot production is provided by Malatya province. There are many pests and diseases affecting apricots in addition to climatic factors such as frost and hail. Partial shoot drying and sudden drying in apricot orchards have increased in recent years. In this study, it was aimed to determine the chemical control possibilities of Neoscytalidium dimidiatum (Penz.) Crous &Slippers, which is one of the fungal agents that cause partial branch drying and sudden drying in apricot orchards in Malatya province by in vitro studies. For this purpose, Kale 4-C isolate of N. dimidiatum collected from Malatya in a previous study was used. Mycelial growth of N. dimidiatum Kale 4-C isolate in PDA were observed in 0 (control), 0.01, 0.03, 0.1, 1, 3, 10, 30 ve 100 μg/mL concentrations of Azoxystrobin 250g/L, Trifloxystrobin %50, Tebuconazole %25, Floupyram 200 g/L+Tebuconazole 200 g/L, Cyprodinil+Fludioxonil %37.5+25, %70 Thiophanate-Methyl and 400 g/L Phosphorous acide fungicides applications. As a result of the study, Floupyram 200g/L+Tebuconazole 200g/L ve Cyprodinil+Fludioxonil %37.5+25 fungicides effectively inhibited the mycelial growth of the fungus.

First Report of Neoscytalidium dimidiatum Causing Stem Canker on Dragon fruit (Hylocereus spp.) in Puerto Rico.

Dragon fruit or pitahaya (Hylocereus spp.) is a tropical fruit belonging to the Cactaceae. It is native to Central and South America and commercially grown in the United States in southern California, south Florida and Puerto Rico. During a disease survey from April to June 2020, stem canker was observed in greenhouses and commercial orchards located in Mayaguez and San Sebastian, Puerto Rico with an incidence of 80%. Diseased cladodes (stems) of 1 mm2 tissue sections of 23 pitahaya varieties (NOI-13, NOI-14, NOI-16, N97-15, N97-17, N97-18, N97-20, N97-22, American Beauty, Cosmic Charlie, Halley's comet, Purple Haze, Alice, Bloody Mary, Dark Star, David Bowie, Delight, Makisupa, Red Jaina, Soul Kitchen, Vietnamese Jaina, Neitzel and Lisa) were disinfested with 70% ethanol, rinsed with double distilled water and plated on potato dextrose agar (PDA) amended with 60 mg/L streptomycin. Three isolates (17B-173-T3, 12C-118-T1 and 13B-131-T2) of Neoscytalidium dimidiatum (syn. N. hyalinum) were identified using taxonomic keys (Crous et al., 2006) and sequencing of the internal transcribed spacer (ITS) with primers ITS5 and ITS4 (White et al. 1990) and translation elongation factor 1 alpha (TEF1-α) with primers EF1-728F and EF1-986R (Carbone and Kohn, 1999). Sequences were compared using the BLASTn tool with N. dimidiatum deposited in NCBI GenBank. In PDA, colonies of N. dimidiatum were initially powdery white and turned grayish-black with age. Arthroconidia (n=50) were dark brown, disarticulating, truncate or cylindrical at the base, thick-walled with 0 to 1 septum, averaging 9.1 X 5.5um in length. GenBank accession numbers of N. dimidiatum DNA sequences were MT921260, MT921261 and MT921262 for ITS and MT920898, MT920899 and MT920900 for TEF1-α. Sequences were 99-100% identical with Ex-isotype CBS145.78 accession numbers KF531816 for ITS and KF531795 for TEF1-α. Pathogenicity tests were conducted on 12 healthy dragon fruit plants of 1.5 years old using three non-detached cladodes per plant. Cladodes were inoculated with 5mm mycelial plugs from 8-day-old pure cultures grown on PDA. Three healthy dragon fruit plants were used as controls and were inoculated with PDA plugs only. The experiment was repeated once. Twenty days after inoculations (DAI), isolates of N. dimidiatum caused stem canker on dragon fruit plants. For all isolates, sunken orange spots averaged 3 X 2 mm in length at 8 DAI. Necrotic blotches with chlorotic halos averaged 10 X 15 mm at 14 DAI; stem cankers with water-soaked tissue were observed at 20 DAI, and arthroconidia and black pycnidia on dry stem cankers at 30 DAI. Untreated controls had no symptoms of stem canker, and no fungi were isolated from tissue. Neoscytalidium dimidiatum has been reported to cause stem canker on Hylocereus spp. in China, Florida, Israel, Malaysia and Taiwan (Chuang et al. 2012; Lan et al., 2012; Ezra et al., 2013; Sanahuja et al., 2016). To our knowledge, this is the first report of N. dimidiatum causing stem canker on dragon fruit in Puerto Rico.

First report of postharvest stem end rot disease on MD2 pineapple fruits caused by Neoscytalidium dimidiatum in Malaysia.

Repeated sampling conducted from December 2019 to March 2020, and fruit of pineapple (Ananas comosus) var MD2 showing early stem end rot symptoms including brown and rotten fruit skin near the stem end region (Fig.1Aa) or darker skin with black discoloration (Fig.1Ab) indicated a consistent fungal infection. The samples (30 fruits from each location) were collected from store houses in three farmer fields with 60% disease incidence in Serdang, (3.0220oN,101.7055oE), Selangor, West Malaysia. The pulp of infected fruits appeared watery with characteristic spoilage odour. Symptomatic necrotic tissues from stem end region and skin were cut in to pieces (1x1cm), surface sterilized and plated onto potato dextrose agar amended aseptically with 0.5 g L-1 streptomycin sulphate. The plates were incubated at room temperature (28±2oC) in natural light conditions. Five days old cultures were light grey in colour and gradually turned dark brown to black with dense deeply tufted, mycelium as the culture aged (Fig.1B, C). Conidial morphology was observed using compound microscope (Olympus model BX-50F4, Tokyo, Japan) equipped with Dino-Eye. Branched mycelia with 0-1 septate arthospores were evident in 14 days old cultures (Fig.1D). Measured arthroconidia (5 to10x3 to 4.5µm) were ellipsoid to ovoid or round shaped, hyaline with an acutely rounded apex, truncate base, initially aseptate (Fig.1E) and arranged as chain at maturity (Fig.1F). The pathogen was identified through PCR amplification of the internal transcribed spacer (ITS) region using ITS1 and ITS4 primers (White et al., 1990) and BLASTn homology search as Neoscytalidium dimidiatum based on 100% similarity to a reference sequence (accession number KJ648577) that was previously deposited (Mohd et al.,2013). The sequence was deposited in Gen Bank ( accession number MW082810). Pathogenicity test was performed using the mycelial plug inoculation method and repeated twice with five replicates. Healthy MD2 pineapple fruits were surface sterilized with 1% NaOCl solution for15 min. followed by washing with sterilized distilled water. One centimeter diameter PDA plug at the margin of actively growing seven days old cultures were inserted in each of two inoculation wounds made on the skin and stem end of each fruit then the wounds were wrapped with moist cotton wool. Non-colonized PDA plugs were used to inoculate the control fruits. Fruits were incubated under 85% RH at room temperature. Five days after inoculation, the fruits showed similar dark necrotic discoloration and confirmed as N.dimidiatum by PCR (Fig.1G). The Koch postulates were fulfilled by inoculation and re-isolation of the fungal pathogen. This pathogen has also been reported previously to cause economic losses on a number of other hosts, such as pitayah fruits in Israel and Malaysia (Erza et al., 2013; Mohd et al., 2013)) and almond in California (Mohomed et al., 2018). To our knowledge this is the first report of N. dimidiatum causing postharvest stem end rot on MD2 pineapple in Malaysia. It may have the possibility to develop postharvest economic losses to pineapple industry, if severely affected fruits with high population of the pathogen left unattended in store houses.

Fungal Pathogens Associated With Canker Diseases of Almond in California.

Almond canker diseases are destructive and can reduce the yield as well as the lifespan of almond orchards. These diseases may affect the trunk and branches of both young and mature trees and can result in tree death soon after orchard establishment in severe cases. Between 2015 and 2018, 70 almond orchards were visited throughout the Central Valley of California upon requests from farm advisors for canker disease diagnosis. Two major canker diseases were identified, including Botryosphaeriaceae cankers and Ceratocystis canker. In addition, five less prevalent canker diseases were identified, including Cytospora, Eutypa, Diaporthe, Collophorina, and Pallidophorina canker. Seventy-four fungal isolates were selected for multilocus phylogenetic analyses of internal transcribed spacer region ITS1-5.8S-ITS2 and part of the translation elongation factor 1-α, β-tubulin, and glyceraldehyde 3-phosphate dehydrogenase gene sequences; 27 species were identified, including 12 Botryosphaeriaceae species, Ceratocystis destructans, five Cytospora species, Collophorina hispanica, four Diaporthe species, two Diatrype species, Eutypa lata, and Pallidophorina paarla. The most frequently isolated species were Ceratocystis destructans, Neoscytalidium dimidiatum, and Cytospora californica. Pathogenicity experiments on almond cultivar Nonpareil revealed that Neofusicoccum parvum, Neofusicoccum arbuti, and Neofusicoccum mediterraneum were the most virulent. Botryosphaeriaceae cankers were predominantly found in young orchards and symptoms were most prevalent on the trunks of trees. Ceratocystis canker was most commonly found in mature orchards and associated with symptoms found on trunks or large scaffold branches. This study provides a thorough examination of the diversity and pathogenicity of fungal pathogens associated with branch and trunk cankers of almond in California.

Further Investigation on Limb Dieback of Fig (Ficus carica) Caused by Neoscytalidium dimidiatum in California.

Fig limb dieback is a cosmopolitan disease caused by Neoscytalidium dimidiatum (Botryosphaeriaceae), characterized by branch and shoot cankers, discoloration of woody tissues, and dieback. The present study investigated the etiology of the disease in California that seems to have become prevalent among fig orchards in the last several years. During orchard surveys in Fresno, Kern, and Madera Counties over 3 years, we isolated consistently and evaluated the pathogenicity of N. dimidiatum under laboratory and field conditions. The effect of summer and winter pruning on the disease severity and the effects of different environmental and mechanical stresses, such as sunburn and wounding by mallets, were assayed. In addition, the susceptibility of six different cultivars and the effects of eradicating cankered shoots from the fig trees as a method to combat the spread of the disease were studied. Pathogenicity tests demonstrated that N. dimidiatum induces cankers on fig, mainly on wounded shoots. Results from the remaining experiments revealed that summer infection leads to more severe canker lesions than those induced by winter infection and that stressed shoots are more susceptible to infection than nonstressed shoots. 'Brown Turkey', 'Conadria', and 'Calimyrna' cultivars (all nonpersistent figs, i.e., needing pollination for fruit development) were less susceptible than the more susceptible 'Kadota', 'Sierra', and 'Black Mission' (all persistent figs, i.e., not needing pollination for fruit development). Canker removal from the orchard seems to be a good agronomic practice to avoid the spread of disease.

First Report of Black Spot Caused by Neoscytalidium dimidiatum on Sisal in Guangxi, China.

Sisal (Agave sisalana Perrine) is an important hard fiber crop that is widely planted in Guangxi, Guangdong, Hainan, Yunnan, and Fujian provinces, China. In July 2019, a new leaf disease of sisal with a disease incident of about 36% was found in Guangxi (Fig.1a~d). The oval or circular black lesions were 2.3 cm to 15.9 cm in length and 1.6 cm to 5.5 cm in width on both sides of the diseased leaves. The central part of the lesions was slightly hollow. The lesions continuously enlarged and ultimately penetrated the leaves. Reddish brown and dark mucus was secreted from the lesions. The junction of lesions and healthy parts was reddish brown to yellow. The diseased leaf fiber and mesophyll tissues were reddish brown and necrotic. Fresh leaf yield was reduced about 30% by the disease, and fiber quality was significantly compromised every year in Guangxi. Six kinds of fungi distinguished by their morphology, size and color of the colonies were isolated from diseased leaf tissues of 60 sisal plants sampled from five different farms in Guangxi. Isolate JMHB1 was isolated at a rate of 95.67%. The isolate JMHB1 was initially white with dense and hairy aerial mycelium, gradually turning dark grey to olive green on PDA (Fig. 2). Conidia, arthrospores, and chlamydospores were observed on PDA in culture (Fig. 3). The conidia formed arthric chains, disarticulating, cylindrical-truncate, oblong-obtuse to doliiform, colorless and transparent, zero- to one-septate, and averaging 4.4 to 13.8 µm × 2.2 to 5.6 µm (n=100). Arthrospores were short columnar, pigmented and transparent, single or formed arthric chains, averaging 5.5 to 17.9 µm × 2.1 to 3.5 µm (n=100). Chlamydospores were dark brown, round or oval, averaging 4.5 to 9.6 µm × 4.5 to 8.6 µm (n=100). Pathogenicity testing was conducted by inoculating 3-year-old healthy sisal plants with PDA plugs (5 × 5 mm) on which the fungus had grown for 5 days. Nine healthy plants were wounded on the leaves with a sterile needle, and mycelial plugs were placed on the wounds, covered with sterile moist cotton, and wrapped with parafilm. Nine control plants were wounded and treated with PDA plugs as the negative control. The test was repeated three times. All treated plants were kept in a greenhouse at ~28 ℃ and 40% RH. After 5 days, only leaves inoculated with isolate JMHB1 showed lesions similar to symptoms observed in the field (Fig.1e~f). The fungus was re-isolated from all nine diseased plants, and no symptoms were observed on the leaves of control plants. Molecular identification of the fungus was made by PCR amplification of the internal transcribed spacer (ITS) region of rDNA, EF1-α gene and β-tubulin gene using primers ITS1/ITS4 (White et al. 1990), EFl-728F/EF1-986R (Carbone and Kohn 1999), TUB2Fd/TUB4Rd (Aveskamp et al. 2009) respectively. The ITS (MT705646), EF1-α (MT733516) and β-tubulin (MT773603) sequences of JMHB1 were similar to the ITS (AY819727), EF1-α (EU144063) and β-tubulin (KF531800) sequences of the epitype of Neoscytalidium dimidiatum (CBS 499.66) with 100%, 99.65% and 99.02% identity, respectively. Based on pathogenicity testing, morphological characteristics, and molecular identification, the pathogen of sisal causing black spot was identified as N. dimidiatum (Penz.) Crous & Slippers (Crous et al. 2006). To our knowledge, this is the first report of black spot caused by N. dimidiatum on sisal in China. Sisal is the main economic crop in arid and semi-arid areas that is widely planted in several provinces of southern China. The serious occurrence of the disease caused by N. dimidiatum has greatly affected the development of sisal industry and local economic income in China. Identification of the pathogen of the disease is of great significance to guide disease control, increase farmers' income and promote the development of sisal industry.

Identification of Botryosphaeriaceae associated with stem-end rot of mango (Mangifera indica L.) in Malaysia.

To identify Botryosphaeriaceae fungal species that are associated with stem-end rot of mango, and to study their pathogenicity on mango fruit. Based on the sequences of internal transcribed spacer (ITS), TEF1-α and β-tubulin, as well as on the phylogenetic analysis of combined sequences, four species of Lasiodiplodia (L. theobromae,L. pseudotheobromae, L. iranensis, L. mahajangana) and two species of Neofusicoccum (N. ribis, N. parvum) were identified. Pseudofusicoccum violaceum, Neoscytalidium dimidiatum and three species of Botryosphaeria (B. scharifii, B. dothidea, B. ramosa) were identified based on sequences of ITS and TEF1-α. Pathogenicity test of selected isolates were tested on Chok Anan, Waterlily and Falan mango cultivars. Generally, all species were observed to be pathogenic on the three tested mango cultivars on wounded fruits, except for N. ribis and N. parvum, which were pathogenic on both wounded and unwounded fruits. However, N. ribis was only pathogenic on cultivar Falan, whereas B. ramosa were pathogenic on cultivars Waterlily and Falan. Eleven species of Botryosphaeriaceae were associated with mango stem-end rot in Malaysia. To the best of our knowledge, four species, namely L. mahajangana, B. ramosa, N. ribis and P. violaceum are the first recorded Botryosphaeriaceae fungi associated with stem end rot of mango. The identification of Botryosphaeriaceae fungi is important to establish suitable control measures and quarantine requirements. Many species have a wide host range, which means that there is a possibility of cross infection from other infected plants.

Development and Validation of an In-House Library for Filamentous Fungi Identification by MALDI-TOF MS in a Clinical Laboratory in Medellin (Colombia)

Identification of filamentous fungi by conventional phenotypic methods are time-consuming, and a correct identification at the species level is prone to errors. Therefore, a more accurate and faster time-to-results, and cost-effective technique, is required, such as the Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry (MALDI-TOF MS). In this study, we describe the development of an in-house spectra library for the identification of filamentous fungi frequently isolated from patients with infections. An in-house spectra library was constructed using 14 reference strains grown in solid medium. Clinical isolates were identified either by the in-house spectra library or the Biotyper commercial library from Bruker Daltonics. Fungal identification was carried following the Biotyper’s established scores: ≤1.699: not reliably identified (NRI); 1.700–1.999: genus-level; ≥2.000: species-level. Clinical isolates were identified, with the in-house library, at species- and genus-level at 88.70% (55) and 3.22% (2), respectively. While 4.80% (3) was NRI and 3.22% (2) was discrepant concerning sequencing. On the contrary, identification up to species and genus-level with the commercial library was 44.44% (16) and 22.22% (8), respectively. NRI and the discrepancy was 30.55% (11) and 2.77% (1), respectively. For the reaming 26 isolates, 16 from Neoscytalidium dimidiatum and 10 from Sporothrix spp., respectively, the absence of spectrum and the specific spectra within the Sporothrix complex in the commercial library resulted in the inability to obtain an identification. In conclusion, the current results advocate the importance that each clinical microbiological laboratory needs to develop an ad hoc library associated with the MALDI-TOF MS fungal identification to overcome the limitations of the available commercial libraries.

First Report of Neoscytalidium dimidiatum Causing Fruit Rot on Guava (Psidium guajava L.) in Malaysia.

Guava (Psidium guajava L.) is an economically important fruit crop in Malaysia with annual production of 67,087 tons in 2018 (FAO 2018). In February 2019, fruit rot symptoms were observed postharvest on approximately 30% of guava cv. Lohan collected from a commercial orchard in the Rawang district (3°23'22.8"N 101°26'55.7"E) of Selangor province, Malaysia. Symptoms on the fruits appeared as small, circular brown spots (ranging 5 to 20 mm in diameter) that coalesced and rapidly expanded to cover the entire fruit. Severely infected fruits became soft and rotted. Ten diseased guava fruits were collected from the sampling location. Small pieces (5x5x5 mm) of symptomatic fruit tissues were excised from the lesion margin, surface-sterilized with 0.5% sodium hypochlorite (NaOCl) for 1 min, rinsed twice with sterile distilled water, plated on potato dextrose agar (PDA) and incubated at 25 °C for 5 days. A Scytalidium-like fungus was consistently isolated from symptomatic tissues on PDA after 4 days. For morphological identification, single-spore cultures were grown on PDA at 25 °C and a representative isolate LB1 was characterized further. The fungal colonies were initially white, powdery, and later turned grayish-black with the onset of sporulation. The mycelia were branched with septa, pigmented, and brown in color. Fungal colonies produced dark-brown arthroconidia with thick-walled, 0 to 1-septa, averaged 9 μm x 5 μm (n=20), and cylindrical to oblong in shape. For molecular identification, genomic DNA was extracted from fresh mycelium of isolate LB1 using DNeasy Plant Mini kit (Qiagen, Germantown, MD, USA). The internal transcribed spacer (ITS) region of rDNA and translation elongation factor 1-alpha (TEF1-α) gene were amplified using ITS5/ITS4 (White et al. 1990) and EF1-728F/EF1-986R primer set (Carbone and Kohn 1999), respectively. Both ITS (954 bp) and TEF1-α (412 bp) sequences exhibited 100% identity to Neoscytalidium dimidiatum with GenBank accession numbers FM211432 and MK495414, respectively. The resulting sequences were deposited in GenBank (ITS: Accession no. MT565490; TEF1-α Accession no. MT572846). Based on the morphological and molecular data, the pathogen was identified as N. dimidiatum (Penz.) Crous & Slippers (Crous et al. 2006). A pathogenicity test was conducted on 5 healthy detached mature guava fruits cv. Lohan by wound-inoculating using a sterile needle and pipetting 10-µl of a conidial suspension (1 × 106 conidia/ml) of isolate LB1 to the wound. Five additional fruits were wounded and pipetted 10-µl sterile distilled water to serve as controls. Inoculated fruits were placed in sterilized plastic container and incubated at 25 ± 1 °C, 90% relative humidity with a photoperiod of 12 h, and the experiment was conducted twice. All inoculated fruits developed symptoms as described above 4 to 7 days post-inoculation, while the control fruits remained asymptomatic. N. dimidiatum was re-isolated from all symptomatic tissues confirming Koch's postulates. N. dimidiatum has been reported causing brown spot disease on pitaya (Lan et al. 2012), and stem canker on dragon fruit in Malaysia and Florida (Mohd et al. 2013; Sanahuja et al. 2016) but this is the first report of N. dimidiatum causing postharvest fruit rot on guava in Malaysia. This disease can cause significant postharvest losses to guava production which could lower marketable yield and proper control strategies should be implemented.

Plant performance of enhancing licorice with dual inoculating dark septate endophytes and Trichoderma viride mediated via effects on root development

BackgroundThis study aimed to assess whether licorice (Glycyrrhiza uralensis) can benefit from dual inoculation by Trichoderma viride and dark septate endophytes (DSE) isolated from other medicinal plants.MethodsFirst, we isolated and identified three DSE (Paraboeremia putaminum, Scytalidium lignicola, and Phoma herbarum) and Trichoderma viride from medicinal plants growing in farmland of China. Second, we investigated the influences of these three DSE on the performance of licorice at different T. viride densities (1 × 106, 1 × 107, and 1 × 108 CFU/mL) under sterilised condition in a growth chamber.ResultsThree DSE strains could colonize the roots of licorice, and they established a positive symbiosis with host plants depending on DSE species and T. viride densities. Inoculation of P. putaminum increased the root biomass, length, surface area, and root:shoot ratio. S. lignicola increased the root length, diameter and surface area and decreased the root:shoot ratio. P. herbarum increased the root biomass and surface area. T. viride increased the root biomass, length, and surface area. Structural equation model (SEM) analysis showed that DSE associated with T. viride augmented plant biomass and height, shoot branching, and root surface area. Variations in root morphology and biomass were attributed to differences in DSE species and T. viride density among treatments. P. putaminum or P. herbarum with low- or medium T. viride density and S. lignicola with low- or high T. viride density improved licorice root morphology and biomass.ConclusionsDSE isolated from other medicinal plants enhanced the root growth of licorice plants under different densities T. viride conditions and may also be used to promote the cultivation of medicinal plants.

Emerging Dematiaceous and Hyaline Fungi Causing Keratitis in a Tertiary Care Centre From North India.

To report rare dematiaceous and hyaline fungal pathogens causing fungal keratitis (FK) at our center and review the published literature in this field. In a retrospective review of FK cases from 2005 to 2011, a total of 14 rare pathogens causing isolated cases of FK of a total of 393 isolates were revived from our collection and reconfirmed by using molecular techniques. The in vitro antifungal susceptibility testing (AFST) was performed against a 6-antifungal drug panel by the CLSI microbroth-dilution method. We reviewed the literature on keratitis due to these rare fungi. Two novel fungi, Alternaria tenuissima and Epicoccum nigrum, were reported in addition to 6 dematiaceous (black pigmented) fungi (Acrophialophora fusispora, Chaetomium globosum, Cladophialophora carionii, Nigrospora sphaerica, Papulaspora equi, and Scytalidium lignicola), 5 hyaline (colorless) fungi (Aspergillus tamarii, Fusarium chlamydosporum, Fusarium incarnatum, Fusarium lichenicola, and Fusarium sacchari), and one yeast (Trichosporon asahii). Amphotericin B had good in vitro activity (minimum inhibitory concentration [MIC] ≤1 μg/mL) against most dematiaceous fungi, but not hyaline fungi (MIC ≥1 μg/mL). Natamycin showed variable MIC, itraconazole and voriconazole had good in vitro activity, except in Fusarium species. Alternaria tenuissima and A. fusispora had a very high MIC (≥16 μg/mL) against echinocandins. Literature search revealed 27 FK cases due to F. lichenicola (n = 6), P. equi (n = 5), F. sacchari (n = 4), A. fusispora (n = 3), S. lignicola (n = 2), and others (n = 7), and more than 50% of these were reported from India. Plant fungal pathogens with variable antifungal susceptibility are an emerging cause of human keratitis with predominance of dematiaceous fungi. Identification and antifungal susceptibility testing are important for epidemiology and to optimize therapy and improve the patient outcome.

Agroindustrial waste as ecofriendly and low-cost alternative to production of chitosan from Mucorales fungi and antagonist effect against Fusarium solani (Mart.) Sacco and Scytalidium lignicola Pesante

This study aimed to evaluate the production of fungal chitosan (FuChi) from Mucorales fungi cultivated in a cashew apple juice (CAJ) and cheese whey (CW) mixture, and to determine the growth-inhibitory effect of this biopolymer against Fusarium solani CFF109 and Scytalidium lignicola CMM1098, which cause root rot disease in cassava plants. Cunninghamella phaeospora UCP 1303 and Cunninghamella elegans UCP 1306 showed the highest FuChi production in screening assay, being selected to a CCRD 22 design to analyze the influence of different CAJ and CW concentrations in the increase of FuChi production. All nine Mucorales fungi cultivated in CAJ-CW medium, showing FuChi production in the range of 27.58 (Mucor hiemalis UCP 1309) to 65.40 mg/g (C. elegans UCP 1306). During CCRD 22 design, the highest FuChi production (64.09 mg/g) was achieved by C. elegans UCP 1306 cultivated in medium containing 40% (v/v) of CAJ and 30% (v/v) of CW, presenting 75% deacetylation degree and crystallinity indexes of 41.50%. FuChi at 16000 μg/mL showed a better inhibition against S. lignicola mycelial growth (81.70%) when compared with F. solani (22.13%) and induced alterations in hyphae morphology on both strains. CAJ and CW are promising substrates for FuChi production, and this biopolymer shows antimicrobial effect against F. solani and S. lignicola.

Diversity and pathogenicity of Botryosphaeriaceae species associated with black root rot and stem cutting dry rot in Manihot esculenta in Brazil

Cassava is a starch-rich, high-calorie tuberous root used as human and animal food, as well as an input material in various industries. Black root rot (BRR) and stem cutting dry rot (SCDR) are cassava diseases that compromise product quality and can greatly reduce crop yields. Consequently, the aim of this study was to identify Botryosphaeriaceae species associated with both diseases in areas of the northeast of Brazil and in areas of the Minas Gerais State in the southeast of the country and infer a correlation between them. Fungal species were identified by morphology combined with phylogenetic analyses of partial sequences of translation elongation factor 1 alpha (tef1-α), internal transcribed spacer (ITS) and RNA polymerase subunit II (rpb2). Five species were identified as being associated with both BRR and SCDR (Lasiodiplodia euphorbicola, L. hormozganensis, L. parva, L. theobromae, and Neoscytalidium dimidiatum), while five others were associated only with SCDR (L. brasiliense, L. caatinguensis, L. iraniensis, L. laeliocattleyae, and L. pseudotheobromae). All species identified in this survey were able to induce disease in cassava roots and seedlings. In decreasing order, the most frequently isolated species were L. theobromae, N. dimidiatum, and L. euphorbicola. This is the first report of L. brasiliense, L. caatinguensis, L. hormozganensis, L. iraniensis, L. laeliocattleyae, and L. parva associated with cassava diseases worldwide.

First report of Neoscytalidium dimidiatum causing tuber rot of potato in Turkey

First report of Neoscytalidium dimidiatum causing tuber rot of potato in Turkey

Neoscytalidium dimidiatum causes leaf blight on Sansevieria trifasciata in Brazil

Sansevieria trifasciata plants showing symptoms of leaf blight were observed in Teresina City, Brazil. Based on morphology and the phylogenetic analysis of DNA sequences of the EF-1α and ITS regions, two isolates of Neoscytalidium dimidiatum were identified. Pathogenicity tests showed that the isolates caused leaf blight on S. trifasciata, in addition to Sansevieria cylindrica, and fulfilled Koch’s postulates. To the best of our knowledge, this report is the first to describe N. dimidiatum on Sansevieria in Brazil.