Prof. Emeritus of Plant Pathology, Mansoura Univ., Seed Health Testing for Medicinal Plants is a Must: Case Studies Anise, Fennel, Fenugreek and Black Cumin Seeds Mohamed A. Elwakil Prof. Emeritus of Plant Pathology, Mansoura Univ., El-Mansoura, Egypt, 35516 Web: mwakil.net E-mail: mawakil@mans.edu.eg ICPS 2015, Dubai

Background: Seed –A Horse of Hunger or Source of Life “1985” Dr. Paul Neergaard, The father of Seed-Pathology Science

Most seed-borne fungi have the potential to produce toxins 1st Statement Most seed-borne fungi have the potential to produce toxins e.g: Acremonium strictum Alternaria alternata Aspergillus species (A. flavus, A. parasiticus, A. fumigatus, A. niger, A. ochraceus, A. nidulans, A. tamari and A. versicolor) Chaetomium globosum Cladosporium cladosporides Curvularia lunata

Continue Fusarium species (F. equiseti, F. oxysporum, F. pallidoroseum, F. solani and F. verticillioides) Penicillium citrinum and Penicillium Chrysogenum Stachybotrys chartarum and Stachybotrys atra Verticillium dahliae and Verticillium albo-atrum

For The Primary identification of associated fungi with seeds 2nd Statement: Seed Pathology Science A- Habit characters (H.C.) of seed-borne fungi on seeds (stereoscopic microscopy) For The Primary identification of associated fungi with seeds

Fig. 1: H.C. of Alternaria radicina on anise seed. Examples Fig. 1: H.C. of Alternaria radicina on anise seed.

Fig. 2: H.C. of Fusarium solani on anise seed.

Fig. 3: H.C. of Fusarium equiseti on black cumin seed.

Fig. 4: H.C. of Fusarium oxysporum on fennel seed.

Fig. 5: H.C. of Verticillium dahliae on fenugreek seed.

Identification of most common mycotoxins produced by seed-borne fungi 3rd Statement Identification of most common mycotoxins produced by seed-borne fungi

Mycotoxins tree Ochratoxin A Aflatoxins (B1, B2, G1, G2) Zearalenone Cladosporin Trichothecenes (TCs) Fumonisins B1, B2 Patulin Emodin Satratoxins, F,G,H Citrinin Mycotoxins tree

Hepatotoxicity Teratogenicity Mutagenicity 4th Statement When using medicinal seeds infested or infected with seed-borne fungi able to produce toxins in our daily diets or medications, acute and chronic effects may occur including: Hepatotoxicity Teratogenicity Mutagenicity

Resulting in: Hepatitis Hemorrhage Immunosuppression Hepatic carcinoma Esophageal cancer Kidney failure

5th Statement Example 1 Seed-borne fungi and toxin they produce Cephalosporium spp. Trichoderma spp. Seed-borne fungi and toxin they produce Trichothecenes T-2 Nevalenol Vomitoxin (Deoxynivalenol) Anguidine (Diacetoxyscipenol) Fusarium spp. Verticillium porium Stachybotrys spp. Myrothecium spp. Trichothecium spp. Powerful inhibitors of protein synthesis

Aspergillus parasiticus Example 2 Aflatoxins B1 B2 G1 G2 Aspergillus flavus Aspergillus parasiticus Aflatoxicosis Carcinogenic

Example 3 Aspergillus spp. Ubiquitous Ochratoxins Ochratoxin A Ochratoxicosis Human Carcinogen Ochratoxins Ochratoxin A Aspergillus spp. (A. ochraceus) (A. niger) Penicillium spp. (P. verrucosum) (P. carbonarius) Ubiquitous

Potent estrogenic metabolite Example 4 Estrogenic Syndrome Potent estrogenic metabolite Zearalenone Gibberella spp. Fusarium spp.

Is seed health testing a must? Yes it is 6th Statement Is seed health testing a must? Yes it is Because When one or more fungus of potential to produce toxins are present on or in seeds, serious implications for human health will, for sure, occur. So far Seed-borne fungi should be a component of the integrated commodity management program of the drug industry.

The Present Status 7th Statement Lack of interest from the Drug industry. Lack of Awareness of care given about Medicinal Seed Health Testing.

What are the methods recommended for seed-health? 8th Statement What are the methods recommended for seed-health? The International Seed Testing Association (ISTA) has tests for seeds of different crops i.e. Standard blotter test Deep-freezing test Agar plate test The authors designed an alkaline seed-bed method for detecting the lurked seed-borne fungi.

9th Statement: To absorb this fact, here is a case study Recorded mycotoxin producing fungi on Fenugreek, Anise, Black cumin and Fennel Genera and species Frequency % Fenugreek Anise Black cumin Fennel Acremonium strictum 5 2 5.5 Alternaria alternata 66 91 63 65 A. radicina 50 10 A. tenuis Aspergillus flavipes A. fumigatus 4 15 18 3 A. parasiticus A. candidus A. clavatus 25 6 A. flavus 60 52 48 67 A. nidulans 38 54 32 A. niger 56 74 75 A. ochraceus 53 26 A. oryzae 20 9.5 A. tamarii 31 16 A. terreus 12 14 A. versicolor

Continue Genera and species Frequency % Fenugreek Anise Black cumin Fennel Chaetomium elatum 2.5 3 C. atrobrunneum 5 C. bostrychodes 10 7 C. globosum C. spirale 3.5 Cladosporium cladosporioides 9 12 16 30 C. herbarum 1.5 4.5 C. oxysporum 15 18 C. sphaerospermum 1 C. tenuissimum 4 C. uredinicola 2

Continue Genera and species Frequency % Fenugreek Anise Black cumin Fennel Curvularia lunata 16 6 4 C. pallescens 1 5 C. senegalensis 2 Fusarium equiseti 2.5 30 F. oxysporum 13 31 F. pallidoroseum 32 3 28 F. solani 20 9 19 F. subglutinans 15 F. verticillioides 42 8 56 Mucor circinelloides 5.5 M. hiemalis M. racemosus Myrothecium verrucaria 7

Continue Genera and species Frequency % Fenugreek Anise Black cumin Fennel Penicillium citrinum 18.5 6 2 7 P. chrysogenum 7.5 18 9 25 P. purpurogenum 11 P. aurantiogriseum P. brevicompactum 4 P. corylcphilum 35 12 P. funiculosum 10 P. janthinellum 8 P. notatum 5 P. spinulosum

Continue Genera and species Frequency % Fenugreek Anise Black cumin Fennel Rhizopus arrhizus 3.5 R. oryzae 2.5 10 13 22 R. stolonifer 36 18 4 Stachybotrys atra 3 42 S. chartarum 20 67 6 Trichoderma hamatum 12 T. harzianum 1 2 6.5 T. viride Trichothecium rosum Verticillium albo-atrum 5 19 V. chlamydosporium V. dahliae 9 7 V. nubilum

Our last words Healthy Plant … Healthy Seed Healthy Seed … Healthy People Health People … Healthy Planet

Acknowledgments I would like to thank Dr. Khalid Ghoneem for his efforts in compiling the data of our work. My thanks also dedicated to Dr. Mohamed El-Metwally for his assistance in preparing the presentation. Thanks to Wael Elwakil, Doctor of Plant Medicine Student at University of Florida for his reviews.

Massage from the Glorious Nelson Mandela

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