Morphological and Molecular Identification of Penicillium digitatum Causing Green Mould of Citrus Fruits in Egypt

Nine Penicillium digitatum isolates were isolated from citrus fruits and tested for their pathogenicity on Navel orange fruit (Citrus sinensis L.). In this regard, the isolates exhibited three different levels of virulence. The isolates were identified morphologically using malt extract agar (MEA) and Czapek yeast extract agar (CYA) for colony characteristics, and MEA for microscopic features. Each one of the isolates has the unique features of P. digitatum, especially the largest conidia (6.1-11.9 µm long X 3.2-8 µm width), and phialides (10.1-21.4 µm long X 4-5.1 µm width). In addition, the isolates were identified on a molecular basis using ITS1 and ITS4 primers to confirm morphological identification. Phylogenetic analysis revealed that there was high variance among the isolates, and there were different relations between some of our isolates and other P. digitatum registered strains originating from different countries all over the world. The sequences were submitted to respective GenBank nucleotide databases with accession No. OR198852, OR198853, OR198854, OR198855, OR198856, OR198857, OR198858, OR198859 and OR198860. We also isolated a new P. digitatum (OB15: OR198859) strain with high virulence and rapid sporulatio

Towards unlocking the biocontrol potential of Pichia kudriavzevii for plant fungal diseases: in vitro and in vivo assessments with candidate secreted protein prediction

Abstract Background Plant fungal pathogens cause substantial economic losses through crop yield reduction and post-harvest storage losses. The utilization of biocontrol agents presents a sustainable strategy to manage plant diseases, reducing the reliance on hazardous chemical. Recently, Pichia kudriavzevii has emerged as a promising biocontrol agent because of its capacity to inhibit fungal growth, offering a potential solution for plant disease management. Results Two novel Pichia kudriavzevii strains, Pk_EgyACGEB_O1 and Pk_EgyACGEB_O2, were isolated from olive brine samples. The microscopic characterization of the strains revealed similar structures. However, there were noticeable differences in their visual morphology. Based on their internal transcribed spacer (ITS) DNA sequences, Pk_EgyACGEB_O1 and Pk_EgyACGEB_O2 strains assigned by GenBank IDs MZ507552.1 and MZ507554.1 shared high sequence similarity (~ 99.8% and 99.5%) with P. kudriavzevii, respectively. Both strains were evaluated in vitro against plant pathogenic fungi. The strains revealed the ability to consistently inhibit fungal growth, with Pk_EgyACGEB_O2 showing higher effectiveness. In addition, both P. kudriavzevii strains effectively controlled grey mold disease caused by B. cinerea in golden delicious apples, suggesting their potential as sustainable and eco-friendly biocontrol agents for post-harvest diseases. Based on a comprehensive bioinformatics pipeline, candidate-secreted proteins responsible for the potent antifungal activity of P. kudriavzevii were identified. A total of 59 proteins were identified as common among the P. kudriavzevii CBS573, SD108, and SD129 strains. Approximately 23% of the secreted proteins in the P. kudriavzevii predicted secretome are hydrolases with various activities, including proteases, lipases, glycosidases, phosphatases, esterases, carboxypeptidases, or peptidases. In addition, a set of cell-wall-related proteins was identified, which might enhance the biocontrol activity of P. kudriavzevii by preserving the structure and integrity of the cell wall. A papain inhibitor was also identified and could potentially offer a supplementary defense against plant pathogens. Conclusion Our results revealed the biocontrol capabilities of P. kudriavzevii against plant pathogenic fungi. The research focused on screening novel strains for their ability to inhibit the growth of common pathogens, both in vitro and in vivo. This study shed light on how P. kudriavzevii interacts with fungal pathogens. The findings can help develop effective strategies for managing plant diseases