Ahmed Mahmoud Ismail

Several Colletotrichum species are able to cause anthracnose disease in coffee (Coffea arabica L.) and occur in all coffee production areas worldwide. A planned investigation of coffee plantations was carried out in Southwest Saudi Arabia in October, November, and December 2022. Various patterns of symptoms were observed in all 23 surveyed coffee plantations due to unknown causal agents. Isolation from symptomatic fresh samples was performed on a PDA medium supplemented with streptomycin sulfate (300 mg L−1) and copper hydroxide (42.5 mg L−1). Twenty-seven pure isolates of Colletotrichum-like fungi were obtained using a spore suspension method. The taxonomic placements of Colletotrichum-like fungi were performed based on the sequence dataset of multi-loci of internal transcribed spacer region rDNA (ITS), chitin synthase I (CHS-1), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), actin (ACT), β-tubulin (TUB2), and partial mating type (Mat1–2) (ApMat) genes. The novel species are described in detail, including comprehensive morphological characteristics and colored illustrations. The pathogenicity of the isolated Colletotrichum species was assessed on detached coffee leaves as well as green and red fruit under laboratory conditions. The multi-locus phylogenetic analyses of the six-loci, ITS, ACT, CHS-1, TUB2, GAPDH and ApMat, revealed that 25 isolates were allocated within the C. gloeosporioides complex, while the remaining two isolates were assigned to the C. boninense complex. Six species were recognized, four of them, C. aeschynomenes, C. siamense, C. phyllanthi, and C. karstii, had been previously described. Based on molecular analyses and morphological examination comparisons, C. saudianum and C. coffeae-arabicae represent novel members within the C. gloeosporioides complex. Pathogenicity investigation confirmed that the Colletotrichum species could induce disease in coffee leaves as well as green and red fruits with variations. Based on the available literature and research, this is the first documentation for C. aeschynomenes, C. siamense, C. karstii, C. phyllanthi, C. saudianum, and C. coffeae-arabicae to cause anthracnose on coffee in Saudi Arabia.

The cucumber (Cucumis sativa L.) is often subjected to several fungal diseases. Rhizoctonia solani-induced cucumber damping-off and root rot are the most common diseases reported from the commercial greenhouses of the eastern area of Saudi Arabia. The objective of the current study is to explore the antagonistic activity of four Trichoderma species against R. solani in vitro and in vivo. Ten R. solani isolates (eight belonging to AG-4 and two belonging to AG-A and AG2-1) were studied. AG4 isolates were pathogenic to cucumber plants, while AG-A and AG2-1 isolates were non-pathogenic. Seven isolates of Trichoderma spp., named T. hamatum KSATR8, T. harzianum (KSATR9 and KSATR10), T. asperellum (KSATR11, KSATC, and KSAT1E), and T. longibrachiatum KSATS were isolated, and the identities of both R. solani and Trichoderma isolates were confirmed based on the phylogenetic analysis of the DNA sequence of the ITS region. The dual culture findings indicated that T. asperellum KSATC and KSAT1E exhibited the most significant inhibitory activities against R. solani, with values of 79.33 and 70.89%, respectively. Scanning electron microscope (SEM) images showed a considerable degradation in the cell wall and collapsing of R. solani hyphae by all Trichoderma species. Under greenhouse conditions, the application of T. asperellum KSATC and KSAT1E at concentrations of 2 × 108 conidia/mL revealed a reduction in root rot and damping-off incidence percentages with values that did not reveal a significant (p < 0.05) difference from those of Rizolex-T fungicide. Nevertheless, the efficacy of the fungicide attained 86.67%, being higher than that of T. asperellum KSATC, which reached 80%. Trichoderma asperellum KSATC and KSAT1E were the greatest in increasing peroxidase, catalase, and chitinase enzymes activities in cucumber plants. Conversely, a significant (p < 0.05) elevation in polyphenol oxidase enzyme (0.762 and 0.97 U/g FW) and total phenol content (0.55 and 0.62 mg/g FW) was recorded in cucumber plants treated with T. harzianum KSATR9 and KSATR10, respectively. The statistical analysis results displayed no considerable variations among cucumber plants regarding total chlorophyll content as a response to treatments with Trichoderma species and fungicides. Therefore, we endorse using T. asperellum KSATC and KSAT1E as an alternative to fungicides to manage root rot and damping-off in cucumbers.

The Alternaria species are considered to produce a plethora of several mycotoxins constituting a risk factor for both human and animal health. This work aimed mainly to explore the cytotoxicity of a combined mixture of altenuene (ALT), alternariol (AOH), tenuazonic acid (TeA), and altenuisol (AS) toxins produced by pathogenic A. alternata toward human oral epithelial cells (PCS-200-014), lung fibroblast cells (WI-38), and male albino rats. The sequencing of the multi-locus, RNA polymerase second largest subunit (rpb2), glyceraldehyde-3-phosphate dehydrogenase (gapdh), and Alternaria major allergen gene (Alt a 1) was performed to infer relationships among isolated Alternaria species. The phylogenetic analysis of gapdh, rpb2, and Alt-a 1 sequence data indicated that all isolates resided in A. alternata. The pathogenic potentiality of A. alternata was investigated on tomato plants cv. super strain B under greenhouse conditions, and all isolates were pathogenic to tomato plants, with significant (p < 0.05) variations. The ability of A. alternata isolates to produce mycotoxins was also explored using high-performance liquid chromatography (HPLC). All tested isolates were able to produce at least one of the assessed mycotoxins—ALT, AOH, TeA, and AS—and ALT was reported as the dominant mycotoxin, produced by 80% of A. alternata isolates. The cytotoxic properties of the combined mixture of ALT, AOH, TeA, and AS at concentrations of 31.25, 62.50, 125, 250, and 500 µg/mL were assessed via the MTT assay method after exposure for 24 h versus the control. The treatment of both cell lines with combined mixtures of ALT, AOH, TeA, and AS showed a dose-dependent decrease in cell viability. The highest concentrations tested at 62.50, 125, 250, and 500 µg/mL significantly decreased cell viability and caused cell damage compared to the lowest concentration of 31.25 µg/mL and the control. The cytotoxicity and genotoxicity of the combined mixtures of ALT, AOH, TeA, and AS on male albino rats were also investigated via the gene expression of (TNF-α) and using hematological (CBC), chemical (alanine aminotransferase (ALT), aspartate aminotransferase (AST) and urea and creatinine), and histopathological analyses. A marked increase was observed in the levels of ALT, AST, urea and creatinine, TNF-α gene expression, red blood cells (RBCs), white blood cells (WBCs), hemoglobin (Hb), and packed cell volume % (PCV) after 28 days of exposure relative to the untreated control. Pathological alterations were also observed in the liver and kidney tissues of rats. Conclusively, this work provides a new understanding on the cytotoxicity and genotoxicity of mycotoxins of pathogenic A. alternata from tomatoes.