Mycelial Colonies Research Articles

Image Analysis Characterizes Phenotypic Variation in the Growth of Mushroom-Forming Fungus Schizophyllum commune.

Schizophyllum commune, a common wood-decay mushroom known for its extremely high genetic variation and as a rare cause of human respiratory diseases, could be a promising model fungus contributing to both biology and medicine. To better understand its phenotypic variation, we developed an image analysis system that quantifies morphological and physiological traits of mycelial colonies in Petri dishes. This study evaluated growth of six wild and one clinical isolates of Japanese S. commune, subjected to different temperatures and glucose concentrations, including a condition mimicking the human respiratory environment. Our analysis revealed that combinations of two growth indices, area and whiteness, and profiling by clustering algorithms highlighted strain-specific responses. For example, the clinical isolate was the whitest under the respiratory-like condition. We also found that the growth rate was strongly determined by glucose concentration, while the effects of temperature on growth varied among the strains, suggesting that while glucose preference is common in this species, responses to temperature differ between strains. This system showed sufficient sensitivity to detect variation in mycelial growth. Our study provides a key to unraveling morphological and physiological traits behind the high polymorphisms in S. commune, including the ability to colonize the human respiratory tract.

First Report of Bulb Rot Caused by Fusarium avenaceum on Fritillaria taipaiensis P. Y. Li in China.

Taibai Beimu (Fritillaria taipaiensis P. Y. Li) is a traditional Chinese herb renowned for its use in alleviating respiratory ailments. In June of 2022 and 2023, an incidence 10 to 20% of foliage symptoms, including yellowing and wilting, was observed in a 10-hectare Taibai Beimu field located in Shangluo, Shaanxi Province (33.64° N, 106.72° E). The middle and stem-adjacent areas of the infected bulbs had brown to black lesions that rotted entirely in severe cases. Fifty symptomatic bulbs were cut into 4 × 4 mm pieces at the junction of the diseased and healthy parts. These pieces were sterilized in 2% sodium hypochlorite for 2 min, followed by 75% ethanol for 30 s, and then rinsed with sterile water. The segments were placed on potato dextrose agar (PDA) and incubated at 28℃ for 5 days in darkness. The morphological characteristics were observed on PDA and synthetic low-nutrient agar (SNA). Sixteen isolates showing similar morphological characteristics were obtained. Representative isolate FTC1 was selected for further analysis. After five days, the mycelial colony on PDA was covered with tomentose aerial mycelia in white or light red. On SNA, the microconidia were oval or rod-shaped, measuring 5.9 to 11.0 × 1.4 to 2.9 µm (average 8.5 × 2.2 µm, n = 50), while the macroconidia were falcate, slender, and curved with an elongated apical cell, usually 1 to 3 septa and ranged from 22.0 to 78.2 × 1.4 to 2.9 µm (average 50.2 × 2.4 µm, n = 50). The morphology was consistent with a previous description of Fusarium avenaceum (Sun et al. 2022). The internal transcribed spacer (ITS) region, translation elongation factor 1-α (EF-1α), RNA polymerase II largest subunit (RPB1), and RNA polymerase II second largest subunit (RPB2), were amplified from extracted genomic DNA with the primer pairs ITS1/ITS4 (White et al. 1990), EF1/EF2 (O'Donnell et al. 1998), RPB1U/RPB1R (Ponts et al. 2020), and 5F2/7cR (Reeb et al. 2004), respectively. BLASTn queries of NCBI GenBank revealed a 100%, 99.7%, 99.6% and 99.1% homology with F. avenaceum (MH864972, MZ153160, MW024696 and MZ078939), respectively. The sequences were deposited in GenBank (ITS, PP879197; EF-1α, PQ106669; RPB1, PQ106668; RPB2, PP554893). The phylogenetic analysis of the multilocus sequences of the isolate FTC1 was conducted using the maximum likelihood method with MEGA11.0 (Tamura et al. 2007). The isolate FTC1 was grouped with F. avenaceum strains NRRL 54939, NRRL 53729 and NRRL 26911; therefore, the isolate FTC1 was identified as F. avenaceum. To fulfill Koch's postulates, fresh FTC1 hyphae were inoculated in 250 ml of potato dextrose broth (PDB) on a shaker at 180 rpm and 28℃ for three days. The spores were filtered through four layers of gauze and adjusted to 1 × 106 spores/ml with sterilized water. Five healthy five-year-old plants were inoculated with 20 ml of spore suspensions, while PDB was used as a control. The inoculation involved pouring the spore suspension over a bulb and covering it with sterile nutrient soil. All plants were grown in sterile nutrient soil in artificial climate chambers (MRC-1100C, Ningbo Prandt Instrument, China) set to 20℃, 80% relative humidity, under a 12-h photoperiod. After 21 days, the plants inoculated with the pathogen exhibited symptoms identical to those observed in the field, while all control plants remained healthy. The pathogen was re-isolated from the symptomatic bulbs, which was confirmed both morphologically and molecularly, as described earlier. F. avenaceum has been reported as a pathogen of Chinese medicinal herbs, such as Polygonatum cyrtonema (Xu et al. 2019) and Coptis chinensis (Mei et al. 2021). To our knowledge, this is the first report of F. avenaceum infecting Taibai Beimu in China, posing threats to its production.

Assessing Pathogenicity and Cultural Characteristics of Fusarium oxysporum f. sp. ricini Isolates from Major Castor Growing Regions of India

The present study was undertaken to investigate the pathogenicity and cultural characteristics of 20 isolates of Fusarium oxysporum f. sp. ricini, the causal fungus of wilt disease of castor (Ricinus communis L). Pathogenicity tests on the susceptible cultivar JI-35 revealed that all isolates were pathogenic, with symptoms progressing from yellowing and drooping of leaves to marginal necrosis, wilting and root discoloration. The incubation period varied significantly among the isolates, ranging from 10.0 to 22.5 days, while the per cent disease incidence (PDI) ranged from 51.7% to 100%. Cultural characterization that includes mycelial colour, colony morphology, pigmentation, growth habit and sporulation highlighted significant variability among the isolates of F. oxysporum f. sp. ricini. The colour of mycelium among the isolates ranged from white to various shades of pink and yellow, with growth habits observed as fast, moderate, or slow. Sporulation rates varied from very high to sparse among the isolates. The observed diversity in pathogenicity and cultural traits emphasized the presence of genetic variability in F. oxysporum f. sp. ricini isolates. These results are vital for understanding the pathogen virulence and for developing effective management strategies for wilt disease in castor crop.

The biosynthesis and impacts of cytokinins on growth of the oyster mushroom, Pleurotus ostreatus

ABSTRACT While a lot is known about cytokinins (CKs) and their actions at the molecular and cellular levels in plants, much less is known about the function of CKs in other kingdoms such as fungi. CKs have been detected in a wide range of fungal species where they play roles ranging from enhancing the virulence of phytopathogens to fortifying plant growth when secreted from fungal symbionts. However, the role of CKs where they concern fungal physiology, apart from plant associations, remains largely uncharacterized. Profiling by UHPLC-HRMS/MS (ultrahigh-performance liquid chromatography–high-resolution tandem mass spectrometry) revealed that Pleurotus ostreatus (oyster mushroom) produces CKs in vitro in both liquid and solid cultures. During fungal growth, CK profiling patterns were consistent with previous suggestions that tRNA degradation products might play a role in the physiological development of fungi. It confirms that those products are CKs that act as fungal growth regulators. Moreover, P. ostreatus was shown to respond to exogenous applications of aromatic and isoprenoid CKs, and their effects were dependent on the dose and CK type in a biphasic manner consistent with hormone action. N 6-benzyladenine (BAP), kinetin (KIN), N 6-isopentenyladenine (iP), and trans-zeatin (tZ) bioassays all revealed hormesis-type responses. Accordingly, at low doses, mycelium colony diameter, biomass accumulation, and changes in morphology were stimulated, whereas at high doses only inhibitory effects were observed. Thus, CKs may act as “mycohormones” and consequently have potential for applications in fungal agriculture and medicinal compound production.

Complexes of cultivated microfungi from peatlands in the mountain landscapes of the Sub-Polar Urals

The study concerned the abundance and species composition of cultivated microfungi from peat soils and hollows of flat-palsa bogs in the mountain landscapes of the sub-Polar Urals (the northern part of National Park Yugyd Va). The number of fungi in the studied peat soils varied from 0.4 to 242 thousand CFUs/g a.d.s. with maximum values in the upper layer of live mosses of Bog I – 242±66.2 thousand CFUs/g a.d.s.. In the bog hollows, the abundance of micromycetes had relatively low values, in hollow of Bog I – 28.6±7.1 thousand CFUs/g a.d.s., in hollow of Bog II – 32.9±25.5 thousand CFUs/g a.d.s. The taxonomic list of cultivated micromycetes included 61 species of fungi from 15 genera, two divisions and Mycelia sterilia. The Mucoromycota division was represented by 17 species from the genera Absidia, Actinomucor, Mucor, Mortierella, and Umbelopsis. The majority if cultivated fungi belonged to the division Ascomycota (43 species from 10 genera). The genus Penicillium dominates by species number (21 species). Reasoning from the frequency of occurrence, the structure of the complex of microfungi included rare and random species by 53%. The share of frequent and dominant fungi accounted for 34 and 13%, respectively. The dominating group consisted of Pseudogymnoascus pannorum and sterile mycelium. Frequent species were Mortierella alpina, Mucor hiemalis, Umbelopsis ramanniana, U. vinacea, Penicillium canescens, P. granulatum, P. lividum, P. simplicissimum, P. spinulosum, P. thomii, P. verrucosum, Talaromyces funiculosus. Most abundant in the layer of live mosses were Penicillium spinulosum (17%), P. thomii (18%), Talaromyces funiculosus (19%). In the seasonally thawed peat layers, the following species were highly abundant as Pseudogymnoascus pannorum (11%), Talaromyces funiculosus (14%), and sterile mycelium (16%). In the gley soil horizons, Pseudogymnoascus pannorum (78%) dominated by abundance. Only single colonies of sterile mycelium were found in frozen peat layers.

Mushrooms grown in high-altitude soil exhibiting distinct alterations in growth, biochemical composition and antioxidant potential

Functional foods like mushrooms are high in secondary metabolites and bioactive chemicals with anti-inflammatory, antibacterial, antidiabetic, immune-stimulating and other health-promoting qualities. This study investigates four mushroom varieties including Cordyceps militaris, Pleurotus ostreatus, Pleurotus djamor and Agaricus bisporus, grown on Indian soil and natural environment for measurement of phenol and flavonoid content, along with their antioxidant properties and biochemical makeup. They had a large amount of phenolic and flavonoid content and high DPPH, H2O2 and NO scavenging inhibition percentages, demonstrating strong antioxidant potential. These mushroom extracts also contained a sizable amount of proline and nutritional content like carbohydrate, protein, lipids and fiber. Among all the mushrooms, the biochemical profile was significantly highest. Significant variations were also seen in the amounts of moisture and ash in each sample. This research aimed to determine how mixing soil with compost substrate affected the yield and growth of mushrooms. The soil had a massive impact on forming more mycelial colonies which were necessary for the entire spawn run and spawning to pin head production and for the maturation of pins. It was discovered through this research that combining soil and compost substrate can aid in the growth of good-quality mushrooms and improve their productivity.

Cross-species transmission of a novel bisegmented orfanplasmovirus in the phytopathogenic fungus Exserohilum rostratum

Mycoviruses have been found in various fungal species across different taxonomic groups, while no viruses have been reported yet in the fungus Exserohilum rostratum. In this study, a novel orfanplasmovirus, namely Exserohilum rostratum orfanplasmovirus 1 (ErOrfV1), was identified in the Exserohilum rostratum strain JZ1 from maize leaf. The complete genome of ErOrfV1 consists of two positive single-stranded RNA segments, encoding an RNA-dependent RNA polymerase and a hypothetical protein with unknown function, respectively. Phylogenetic analysis revealed that ErOrfV1 clusters with other orfanplasmoviruses, forming a distinct phyletic clade. A new family, Orfanplasmoviridae, is proposed to encompass this newly discovered ErOrfV1 and its associated orfanplasmoviruses. ErOrfV1 exhibits effective vertical transmission through conidia, as evidenced by its 100% presence in over 200 single conidium isolates. Moreover, it can be horizontally transmitted to Exserohilum turcicum. Additionally, the infection of ErOrfV1 is cryptic in E. turcicum because there were no significant differences in mycelial growth rate and colony morphology between ErOrfV1-infected and ErOrfV1-free strains. This study represents the inaugural report of a mycovirus in E. rostratum, as well as the first documentation of the biological and transmission characteristics of orfanplasmovirus. These discoveries significantly contribute to our understanding of orfanplasmovirus.

Sensitivity of Globisporangium ultimum to the fungicide metalaxyl is enhanced by the infection with a toti-like mycovirus

In recent years, numerous oomycete mycoviruses have been discovered; however, very few studies have focused on their effects on the host oomycete phenotype. In this study, we investigated the impact of toti-like Pythium ultimum RNA virus 2 (PuRV2) infection on the phytopathogenic soil-borne oomycete Globisporangium ultimum, which serves as a model species for Globisporangium and Pythium, specifically the UOP226 isolate in Japan. We generated a PuRV2-free isogenic line through hyphal tip isolation using high-temperature culture and subsequently compared the phenotypic characteristics and gene expression profiles of UOP226 and the PuRV2-free isogenic line. Our findings revealed that the metalaxyl sensitivity of UOP226 was greater than that of the PuRV2-free isogenic line, whereas the mycelial growth rate and colony morphology remained unchanged in the absence of the fungicide. Furthermore, transcriptome analyses using RNA-seq revealed significant downregulation of ABC-type transporter genes, which are involved in fungicide sensitivity, in UOP226. Our results suggest that PuRV2 infection influences the ecology of G. ultimum in agricultural ecosystems where metalaxyl is applied.

First Report of Fusarium vanettenii Causing Fusarium Root Rot in Fatsia japonica in China

Fatsia japonica plays an important role as a commonly used plant in urban landscaping. From 2022 to 2023, a root rot infestation was observed that caused extensive wilting of Fatsia japonica without leaf shedding and eventual death of the plant, severely reducing the ornamental qualities of the plant as well as the vigor of its growth. Fusarium species were isolated from the roots of the affected plants, exhibiting abundant and dense yellow mycelial colonies that proliferated radially from the center of the Petri dishes. Morphological examinations revealed the presence of falciform macro- and microconidia consistent with Fusarium, as well as chlamydospores characterized by their thick walls. For further identification, the amplification and sequencing of the ITS, TEF1 alpha, and RPB2 alpha genes were performed. Finally, healthy Fatsia japonica plants were inoculated with a spore suspension of the pathogen, to confirm that the disease symptoms were compatible with naturally occurring infection. Fusarium vanettenii was identified as the causative agent of Fatsia japonica root rot. To the best of our knowledge, this is the first report of F. vanettenii causing root rot of Fatsia japonica in China.

Evaluation of Genetic Diversity and Agronomic Traits of Germplasm Resources of Stropharia rugosoannulata

China possesses abundant wild germplasm resources and a wide range of cultivated strains of Stropharia rugosoannulata. However, issues such as genetic diversity and unclear genetic relationships have had an impact on the classification and preservation of germplasm resources, the breeding of new varieties, and the promotion of superior strains. There is an urgent need for genetic diversity analysis and assessment of germplasm resources. In this study, we conducted whole-genome resequencing of 50 cultivated and wild strains collected from various regions across the country. After applying a series of filtering parameters, we obtained 888,536 high-quality Single Nucleotide Polymorphism (SNP) markers. Using these SNP markers, we performed principal component analysis, population structure analysis, and phylogenetic clusters analysis on the 50 strains. Most cultivated strains exhibited high genetic similarity, while significant genetic diversity was observed among wild strains. Based on factors such as marker distribution throughout the entire genome and marker quality, we selected 358 core SNP markers to construct SNP fingerprints. Two-dimensional barcodes were generated for each strain to enable specific identification. Additionally, the mycelial growth rate of strains was evaluated on Potato Dextrose Agar (PDA) and substrate culture media. We also assessed their lignin degradation capability using guaiacol agar plates assay. It was observed that the mycelial growth rate on PDA and substrate culture medium exhibited a significant correlation with the diameter of the mycelial colony on guaiacol agar medium. Additionally, the correlation between the mycelial growth rate on the substrate medium and that on the guaiacol agar plate was stronger than that on PDA medium. This study provided molecular-level identification and assessment of germplasm resources. It clarified the genetic relationships among strains and the characteristics of mycelium growth-related agronomic traits of each strain. This research contributed to the enrichment and development of utilizable germplasm resources and breeding materials for S. rugosoannulata, offering a scientific basis for further research.

Culture media and temperature effects on Neofusicoccum mediterraneum: assessing mycelial growth and colony characteristics

Culture media and temperature effects on Neofusicoccum mediterraneum: assessing mycelial growth and colony characteristics

Antifungal activity and mechanism of palmarosa essential oil against pathogen Botrytis cinerea in the postharvest onions.

Botrytis cinerea is a pathogenic fungus that infests multiple crops, which causes a severe decrease in yield and generates substantial losses in the economy. Palmarosa essential oil (PEO) is a primary aromatic compound extracted from palmarosa that is commonly used for scent, medicine, and flavoring foods due to its diverse bioactive properties. In this study, we explored the antifungal activity and the main mechanism of action of PEO against B. cinerea. In addition, the components and control effects of PEO were also studied. The antifungal assay was tested using the mycelial growth rate method and colony morphology. The constituents of PEO were identified according to gas chromatography/mass spectrometry (GC-MS). The main mechanism of action of PEO was evaluated by measuring representative indicators, which consist of cell contents leakage, excess reactive oxygen species (ROS), and other related indicators. The results indicated that at a concentration of 0.60ml l-1, PEO exhibits strong antifungal activity against B. cinerea. The PEO mainly included 13 compounds, of which citronellol (44.67%), benzyl benzoate (14.66%), and acetyl cedrene (9.63%) might be the main antifungal ingredients. The study elucidated the main mechanism of action of PEO against B. cinerea, which involved the disruption of cell membrane structure, resulting in altered the cell membrane permeability, leakage of cell contents, and accumulation of excess ROS. PEO is a satisfactory biological control agent that inhibits B. cinerea in postharvest onions. PEO (0.60ml l-1) exhibited strong antifungal activity by disrupting the cell membrane structure, altering cell membrane permeability, leading to the cell contents leakage, accumulation of excess ROS and increased level of Malondialdehyde (MDA) compared to the control group.

Cultural Characteristics of Phylloplane Fungi and Leaf Spot Pathogens Isolated from Cajanus cajan

Four different isolates of fungi were discovered from the phylloplane of Arhar. They were recognized and identified to be Fusarium solani, Paecilomyces marquandii, Aspergillus niger and Aspergillus flavus. Similarly, from the diseased leaves of Arhar two pathogens were isolated, characterized and identified to be Curvularia lunata and Sterile. These fungal isolates were cultivated on seven different culture media viz., Czapek’s Dox Agar, Potato Dextrose Agar (PDA), Oat Meal Agar, Rose Bengal Agar, Richard’s Synthetic Agar, Sabouraud’s Dextrose Agar and Malt Extract Agar. Right after seven days of incubation at 250C, the mycelial growth rate, colony character and sporulation patterns of these fungi were evaluated. The type of growth media utilized had a significant impact on the diameter, cultural traits and sporulation of a few selected test fungi. On PDA and Richard’s synthetic agar media, Fusarium solani, Aspergillus niger and Curvularia lunata recorded the superlative radial growth. In PDA and Sabouraud’s dextrose agar Aspergillus flavus and Sterile recorded the highest growth. Paecilomyces marquandii recorded the maximal radial growth in Sabouraud’s dextrose agar.

Identification and Characterization of Endophytic Fungi from Garcinia atroviridis for Potential Antagonistic Against Phytopathogenic, Colletotrichum gloeosporioides

Biological control is referred to as the “use of natural or modified organisms, genes” to minimize the effects of undesirable pests, pathogenic microorganisms, and diseases on plant crops. This measure has become a suitable and safe alternative for chemical fungicides in plant disease management. Endophytic fungi have received much attention as biological control agents against many plant pathogens through antibiosis, parasitism, invading spores, mycelium, and cells of the pathogen, and secreting bioactive metabolites. While the therapeutic properties of Garcinia atroviridis have been studied, the existence of microbial endophytes and their properties is still less documented. In this research, G. atroviridis endophytic fungi were isolated and identified by fungal colony morphology observation combined with the PCR-amplified fungal internal transcribed spacer (ITS) sequence analyses. Fungal endophytes were assessed for their biocontrol potential against Colletotrichum gloeosporioides. In total, 111 endophytic fungal isolates harboring inside the leaf, branch, and fruit of G. atroviridis belonged to 5 different species with 3 different genera and two unidentified genera. All the endophytic fungal species isolated were evaluated using an in vitro dual culture assay against C. gloeosporioides, a common pathogen that causes anthracnose disease. The results of the present study clearly showed that seven species of isolated fungal endophytes were capable of inhibiting the mycelial colony growth of C. gloeosporioides with an inhibition percentage between 54.67% to 87.94%. Among these species, Nigrospora sphaerica recorded the highest PIRG with 87.94%. Our work indicates that endophytic fungi isolated from G. atroviridis have a biocontrol effect on C. gloeosporioides and are expected to be a potential source of bioactive metabolites.

Studi Pertumbuhan Miselia Jamur Merang (Volvariella volvaceae) G2 Isolat FP005 Faperta UNSIKA Pada Beberapa Bahan Alternatif Media In Vitro

Straw mushroom is one of the horticultural commodities that is in great demand by the community. The successful cultivation of straw mushrooms is determined by the quality of the seeds of the molten mushroom. Tissue culture is one of the efforts to get quality seeds. Quality seedlings are seen from the brood of the merang fungus and the quality of pure culture. Making pure culture is still constrained in making in vitro media, especially alternative media materials. This study aims to obtain the best results from the growth of G2 FP005 mycelia on alternative media materials in vitro. This research was conducted at the Biotechnology and Plant Breeding Laboratory, Faculty of Agriculture, Singaperbangsa University, Karawang from April to May 2023. The research method used was a single factor Complete Randomized Design (CRD) with 7 treatments and 5 replications: A (100% PDA); B (100% Charcoal Husk Solution); C (100% Husk Solution); D (80% PDA + 20% Husk Charcoal Solution), E (75% PDA + 25% Husk Solution), F (50% Husk Charcoal Solution + 50% Husk Solution); G (50% PDA + 25% Husk Charcoal Solution + 25% Husk Solution). The effect of the treatment was analyzed using variance and if the F test was significant at 1% level, then to find out which treatment was the best it was continued with the DMRT (Duncan Multiple Range Test) advanced test at the 1% level of significance. The results achieved from this study were that there was a significant effect of using alternative media materials on the growth of the straw mushroom mycelia. Alternative media 80% PDA + 20% rice husk charcoal gave the highest mycelia colony diameter of 7.88 cm and the highest mycelial colony growth rate of 2,878 cm/day. Keywords: Unsika Faperta Isolate, Straw Mushroom, Tissue Culture.

Morphological characterization of Sclerotinia sclerotiorum causing fruit rot of papaya and its management using biopesticides

Papaya is an important fruit crop and susceptible to several post-harvest diseases at every stage of its life cycle. Papaya fruit rot caused by Sclerotinia sclerotiorum, is a serious postharvest disease that results in significant losses both in storage and on the market. Papaya fruit that was gathered from several Lahore fruit marketplaces was the subject of the studies. Based on physical and cultural traits, diseased papaya fruits were separated and identified. The virus developed mycelial colonies on PDA media that were sparse and creamy white. These colonies ultimately became fluffy and light brown. Sclerotia began to appear after three days of mycelial growth, and after seven days, they were fully visible in the shape of rings or distributed across the mycelial surface of the Petri plate. This allowed for the calculation of the frequency of isolated fungi. It was recorded maximum percentage of S. sclerotiorum (58%) was recorded followed by Alternaria alternata (21.5 %) and Aspergillus niger (10.0 %), while Penicillum spp. was least isolated fungi with a mean value of 5.00%. The assessment of different plant extracts including (Cascabela thevetia, Euphorbia milii, Moringa oleifera, Plumeria rubra, Quisqualis indica, Syzygium aromaticum) was done with Agar Well Diffusion Technique under CRD design. The findings showed that of the medicinal plants that were extracted using methanol and ethanol, Moringa oleifera and Syzygium aromaticum showed significant results at higher concentrations and showed maximum zone of inhibition while Cascabela thevetia was least effective against S. sclerotiorum.

First Report of Berkeleyomyces basicola Causing Black Shoot Rot of Raspberry in Nursery Production in the United States and Worldwide.

Commercial production of raspberry (Rubus ideaus) transplants is almost exclusively accomplished through clonal propagation. One system involves forcing young shoots to grow from roots. The shoots are cut and rooted in propagation trays and referred to as tray plants. Sanitation is important during tray plant production as this method carries some risk due to contamination by substrate pathogens. In May 2021, a new disease was observed on raspberry tray plant cuttings at one nursery location in California, and observed again in 2022 and 2023 but at a much lesser extent. Multiple cultivars were affected; however, up to 70% mortality was observed on cv. RH740.1. In less affected cultivars, mortality ranged from 5-20%. Symptoms included chlorotic leaves, lack of rooting, and blackening at the basal end of shoots, followed by death of the cutting. Affected propagation trays had inconsistent foliage and patchy growth. Chains of chlamydospores (two to eight spores in each chain) similar in morphology to Thielaviopsis species (Shew and Meyer 1992) were observed at the cut end of symptomatic tray plants using a microscope. Isolates were retrieved by incubating tissue on surface-disinfested (1% NaOCl) carrot discs in a humid chamber for 5 days until greyish black mycelium was observed (Yarwood 1946). Mycelium was transferred to acidified potato dextrose agar and formed a gray to black compact mycelial colony with both endoconidia and chlamydospores. Endoconidia were catenulate, single-celled with slightly rounded ends, colorless, and 10-20 µm x 3-5 µm in size; dark-colored chlamydospores were 10-15 µm x 5-8 µm in size. The ITS region of isolates 21-006 and 22-024 was amplified with ITS5 and ITS4 primers using a 48°C annealing temperature (White et al. 1990), Sanger sequenced (GenBank accession OQ359100) and yielded 100% match to Berkeleyomyces basicola accession MH855452. Pathogenicity was confirmed by dipping 80 grams of roots of cv. RH740.1 into a suspension of 106 conidia/mL of isolate 21-006 for 15 min. For the non-inoculated control, 80 grams of roots were dipped in water. Roots were then planted into trays of coir (Berger, Watsonville, CA). Six weeks after inoculation, twenty-four shoots were harvested from each treatment, stuck into propagation trays filled with coir and maintained in a humid chamber for 14-days to induce rooting. Tray plants were then harvested and assessed for root development, black basal shoot tips, and presence of chlamydospores. Forty-two percent of cuttings from the inoculated treatment had rotten basal tips and failed to root, in comparison to 8% of the cuttings from the non-inoculated control. Chlamydospores were visualized only on shoots that emerged from inoculated roots and B. basicola was isolated only from cuttings originating from inoculated roots. Post-inoculation isolates were confirmed as B. basicola using methods described above. To our knowledge, this is the first report of B. basicola infecting raspberry. Confirmation of this pathogen on tray plants is significant because of the potential impact this disease may have in commercial nursery production worldwide. In 2021, the value of the harvested raspberry crop in the U.S. totaled $531 M, of which California represented $421 M (USDA 2022). The value of the 2021 crop was highest in the U.S. ($531 M), followed by Russia ($512 M), Spain ($405 M) and Mexico ($332 M) (FAO 2021).

Diagnosis and Biocontrol of Sporisorium scitamineum Associated with Whip Smut Sugarcane

Aim: Whip smut of sugarcane is the most serious and widely spread disease of sugarcane and causes a significant reduction in cane quantity and quality. This work aimed to assess the macroscopic and microscopic characteristics and the variability between isolates of Sporisorium scitamineum causal agent of smut in sugarcane.
 Materials and Methods: The fungus was isolated on Potato Dextrose Agar (PDA) medium. The morphological characters were described based on the growth diameter, texture, and colony colors. The microscopic description focused on the spore’s size from medium and teliospores from whips. Results: The colony appeared with white mycelial cotton and a yeast-shaped mycelial colony. In vitro, control of S. scitamineum isolates with endophytic fungi of Macrophomina phaseolina and Trichoderma viride was studied. The results showed that the two fungi could inhibit the growth of S. scitamineum. A pathogenicity test was carried out on sugarcane cuttings by inoculating the buds with spore suspension of S. scitamineum, which showed a pathogenic effect ranging from 50–100%. Also, other sugarcane cuttings were inoculated by the pathogen and Trichoderma viride as a biocontrol agent that inhibited the pathogen growth completely in four strains.
 Conclusion: Trichoderma viride is the safely biocontrol agent against S. scitamineum.

Óleos essenciais no manejo de Alternaria alternata f. sp. citri em frutos de tangerineira ‘Dancy’

ABSTRACT Tangerines and their hybrids are affected by economically essential diseases. The Alternaria brown spot (Alternaria alternata f. sp. citri) deserves to be highlighted, as it is present in all tangerine production areas. This study aimed to determine the effect of essential oils on A. alternata f. sp. citri. The experiments were carried out in the Laboratories of Phytopathology and Biology and Technology of Post-Harvest of the Federal University of Paraíba, Areia, PB. Three pathogen isolates were used, and ten essential oils from grape, sunflower, eucalyptus, ginger, copaiba, mint, fennel, citronella, clove, and linseed at a concentration of 1%, diluted in potato dextrose agar for in vitro tests. The oils were diluted in distilled water for the fruit test, and the fruits were immersed in the treatments for 5 min. In addition, the fungicide Thiabendazole (400 mL/100L) and sterile distilled water were used as the control treatments. The fruits were inoculated with a conidia suspension of the pathogen (105 conidia/mL). Mycelial Growth Rate Index, colony diameter, spore production and dimensions, fruit disease severity, and enzymatic activity were evaluated. A completely randomized design was used, with 12 treatments and four replications in vivo. The results indicate that the essential oils of mint, eucalyptus, fennel, and citronella were efficient in the in vitro control of the pathogen, ultimately inhibiting the growth of fungal colonies. Eucalyptus oil efficiently managed A. alternata f. sp. citri on ‘Dancy’ tangerine fruits. Essential oils did not influence the enzymatic activity of the fruits.

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

Dragon fruit (Hylocereus spp.) is a tropical perennial plant (Family Cactaceae) that is popular for consumption in India. Originated from Central and South America and currently became popular in India. Hylocereus undatus and H. polyrhizus are the most commonly cultivated species in India. During August to December, 2021, stem cankers were observed in commercial orchards within the Satara, Pune and Solapur districts of Maharashtra. The disease incidence across four orchards was approx. 40% and severity ranged between 30 and 80%. Initial symptoms on infected cladodes were minute, circular, depressed chlorotic spots often with a brick red flecks followed by elevation of the centre of the lesion. Later the lesions turned necrotic and contained black, erumpent pycnidia, followed by chlorosis and stem rot. Twelve diseased cladodes from different orchards were collected and isolations were conducted. Edges of the lesions (5 to10 mm2) were excised and surface sterilized by exposure to 1% sodium hypochlorite (2 min) followed by triple rinsing with sterilized distilled water. Excess moisture was removed with sterilized blotter paper and pieces were plated on potato dextrose agar (PDA) amended with streptomycin sulphate (30 mg/L) for 3 days at 27 ± 2°C with a 12 h photoperiod. All twelve isolation attempts yielded uniform fungal colonies and out of these, purified colonies from each location viz., SLNeo, LNeo, MGNeo and KNeo were selected for etiology study. On PDA, initially white mycelial colonies turned to olive green to grayish with dark gray to black pigmentation. Colony growth was rapid (30 mm/day). Brown, branched septate hyphae fragmented to produce abundant arthroconidia in chains that were hyaline to dark brown, thick walled, 0 to 1 septate, ellipsoid to ovoid (10.1 ± 1.4 × 4.88 ± 1.1 μm), rod (9.0 ± 0.8 μm × 5.3 ± 0.8 μm), round (7.9 ± 1.6 μm) and capsule (14.2 ± 3.1 × 6.1 ± 0.8 μm) shape. Conidia from pycnidia developed on host tissues were aseptate, hyaline and ellipsoid-cylindrical (8.2 ± 0.8 × 2.2 ± 0.3 μm) in shape. Based on morphological characteristics, the isolates were identified as Neoscytalidium dimidiatum (Penz.) Crous & Slippers (Crous et al. 2006). Molecular characterization was done by amplifying the partial internal transcribed spacer (ITS) region, and translation elongation factor 1-α and β-tubulin genes of four isolates using ITS1/ITS4, EF1-728F/EF1-986R and BT2A/BT2B primers (White et al. 1990, Glass and Donaldson 1995, Carbone and Kohn 1999). Sequences were deposited in GenBank accesions (ITS: OM884028, OM884029, OM884030, OM899800; TUB2: OM927962, OM927963, ON099066; TEF1-α: OM927965, OM927966, OM927964, OM984744) showed 99 to 100% identity with the epitype CBS 499.66 accession numbers KF531820 for ITS and KF531798 for TEF1-α and KF531800 for TUB2. N. dimidiatum isolate designated as SLNeo has been deposited at Microbial Type Culture Collection (MTCC), Chandigarh, India with MTCC 13250. Phylogenetic analysis using maximum likelihood (ML) revealed these isolates were clustered with N. dimidiatum clade. To evaluate pathogenicity, stems of one year old dragon fruit plants were wounded with a sterilized needle and inoculated with 7 mm fresh mycelial discs. For each isolate three replications were kept. Non-inoculated controls were plants receiving only non-colonized PDA discs. Inoculated and control plants were kept separately at ambient temperature (35 ± 2°C during day and 22 ± 2°C at night). Symptom started to develop at 2 days after inoculation (DAI) and canker lesions with chlorotic halos and rotting were observed at 15 DAI. No symptoms were observed on the negative control. Pathogenicity testing was repeated twice and the pathogen was re-isolated from symptomatic tissues with 100% re-isolation frequency and found identical with N. dimidiatum. N. dimidiatum has been reported on H. undatus and H. polyrhizus in subtropical and tropical countries worldwide (Chuang et al. 2012; Ezra et al. 2013; Lan et al. 2012; Sanahuja et al. 2016; Serrato-Diaz and Goenaga 2021). To our knowledge, this is the first report of dragon fruit stem canker caused by N. dimidiatum from India. Since the disease poses a major threat to dragon fruit plantations, additional epidemiological studies may assist in developing management strategies.