Endophytic fungi are used as the most common microbial biological control agents (MBCAs) against phytopathogens and are ubiquitous in all plant parts. Most of the fungal species have roles against a variety of plant pathogens. Fungal endophytes provide different services to be used as pathogen control agents, using an important aspect in the form of enhanced plant growth and induced systemic resistance, produce a variety of antifungal secondary metabolites (lipopeptides, antibiotics and enzymes) through colonization, and compete with other pathogenic microorganisms for growth factors (space and nutrients). The purpose of this review is to highlight the biological control potential of fungal species with antifungal properties against different fungal plant pathogens. We focused on the introduction, biology, isolation, identification of endophytic fungi, and their antifungal activity against fungal plant pathogens. The endosymbionts have developed specific genes that exhibited endophytic behavior and demonstrated defensive responses against pathogens such as antibiosis, parasitism, lytic enzyme and competition, siderophore production, and indirect responses by induced systemic resistance (ISR) in the host plant. Finally, different microscopic detection techniques to study microbial interactions (endophytic and pathogenic fungal interactions) in host plants are briefly discussed.

Alternaria alternata is a major spoilage fungus of dairy products and a potential producer of mycotoxins. This study evaluated Pulsed Light (PL) as a non-thermal strategy to control A. alternata FCS26 strain isolated from Italian Caciocavallo cheese and investigated its physiological and transcriptional stress responses. To the best of our knowledge this is the first report on the anti-fungal mechanism of PL against A. alternata. In vitro, treatments with 30 PL (11.46 J cm^-2) and 120 PL (45.84 J cm^-2) differentially inhibited fungal structures: spore germination was reduced by 77% and 83%, respectively, while mycelial growth was inhibited by up to 22% and 37%, respectively. PL treatment also extended the lag phase and decreased cell viability as confirmed by confocal microscopy. Furthermore, PL pronounced oxidative and nitrosative stress, as evidenced by the early accumulation of Reactive Oxygen (ROS) and Nitrogen (RNS) Species and the modulation of antioxidant enzymes. Superoxide dismutase (SOD) and peroxidase (POD) showed sustained enzymatic responses under the 30-pulse treatment, whereas the 120-pulse treatment induced strong early activation followed by late disruption of POD and catalase (CAT) activity, consistent with severe redox imbalance. Reverse transcription-quantitative PCR revealed broad transcriptional modulation, with consistent downregulation of genes encoding polyketide synthase, chitinase, and dedh DEAD/DEAH box helicase-like protein. In situ studies showed that PL significantly reduced fungal severity on Caciocavallo cheese during refrigerated storage. In fact, the application of 120 pulses yielded the strongest effect, decreasing fungal growth severity by about 80% compared to the untreated control. Overall, PL effectively inhibited A. alternata, supporting its potential for fungal control, shelf-life extension and improved dairy safety.

Mold contamination is a major cause of spoilage in ripened cheeses. In this study, 171 fungal isolates were recovered from semi-hard Caciocavallo cheese sampled across three seasons. The number of recovered isolates showed a descriptive decrease from summer to winter. Thirty-two representatives were selected for ITS and/or β-tubulin sequencing after morphotype screening. Among these sequenced representatives, Penicillium commune was the most frequent provisional taxonomic designation (8/32; 25.0%), followed by Aspergillus flavus (3/32; 9.4%). Penicillium echinulatum, Fusarium iranicum, Paecilomyces variotii, and Cladosporium-like isolates each accounted for 2/32 (6.3%). ITS-only assignments in taxonomically difficult genera are treated as provisional closest-match designations rather than definitive species identifications. Essential oils (EOs) from three Salvia rosmarinus Spenn. chemotypes (SR-CV2-23, SR-CV3-23, and SR-CV6-23), oregano cv. Carva (Origanum vulgare L. subsp. viridulum (Martrin-Donos) Nyman × O. vulgare L. subsp. hirtum Ietswaart; Ov-C-23), and Laurus nobilis L. (Ln-23) were chemically characterized and evaluated for their antifungal activity against mold species associated with Caciocavallo cheese spoilage. GC-MS showed that the rosemary EOs were rich in camphor, α-pinene, and 1,8-cineole; Ov-C-23 contained 83.95% carvacrol; and Ln-23 contained mainly 1,8-cineole, linalool, and bornyl acetate. All EOs exhibited concentration-dependent antifungal activity, with spore germination generally more sensitive than mycelial growth. At 0.35% (v/v), SR-CV2-23 showed the broadest observed in-vitro activity, whereas Ov-C-23 produced the highest observed inhibition of A. alternata mycelial growth. CFDA/PI confocal microscopy showed the largest relative shift toward reduced metabolic activity and membrane compromise after SR-CV3-23 treatment. In-vitro assays and cheese-slice experiment used different quantitative endpoints and were therefore not directly compared. Ov-C-23 produced the highest observed reduction in visible colonies in the cheese slice model. These EOs may contribute to integrated mold control strategies, but practical application requires further validation under industrial conditions.