Monograph On

Monograph On Mycotoxigenic Fungi and Mycotoxins in Foods and Feeds with Synopsis of studies done by the authors on mycotoxigenic fungi and mycotoxins in foods and feeds Prepared by Mohamed K. Refai1 and Atef A.Hassan2 1 Profesor of Microbiology, Faculty of Veterinary Medicine, Cairo University, Egypt, e-mail [email protected], 2 Profesor of Mycology and Mycotoxins, Animal Health Research Institute, Dokki, Egypt, e- mail, [email protected] 2013 1 Preface This monograph is designed as a laboratory guide for the food microbiologists to assist in the isolation and identification of common mycotoxigenic foodborne fungi and reporting their respective mycotoxins.We emphasise the fungi which cause food spoilage and mycotoxicosis but also devote space to the fungi commonly encountered in foods and feeds at harvest, and in the food factory. As far as possible, we have kept the monograph simple. The identification keys have been designed for use by microbiologists with little or no prior knowledge of mycology. For identification to genus level, they are based primarily on the cultural and physiological characteristics of fungi grown under a standard set of conditions. The microscopic features of the various fungi become more important when identifying isolates at the species level. Nearly all of the suspected mycotoxigenic species have been illustrated with colony photographs, together with photomicrographs or line drawings. The tables of mycotoxins produced by these fungi were sited briefly as rapid and simple illustration for graduate students.The knowledge about the target organs, systems and clinical symptoms due to toxicosis of each mycotoxin are also available in this monograph. Among the colleagues who helped us to prepare this monograph, we wish to particularly thank Dr. Manal A.Hassan, Chief Researcher of mycology , Animal Health Research Institute, Cairo, Egypt, for her assistant in collection of the papers about the methods of control of mycotoxicosis. We are indebted to Dr. Rasha H. Sayed El-Ahl, Reasercher of mycology, Animal Health Research Institute, Cairo, Egypt, who assisted in literature searches and some cultural and photographic work We wish to express our appreciation to all staff members of Mycology Department, who generously provided facilities, cultures and advice on some of the genera studied. Mohamed K. Refai and Atef A. Hassan November, 2013, Egypt 2 My great respect to the great Disability challenger Prof. Dr. Atef Abdelaziz Hassan When he came to my office 1988 he gave me a paper on which he wrote that he wanted to register for M.V. SC. In the field of mycotoxins and hoped to accept him under my supervision, in spite of his impairment in hearing and talking. I accepted on the spot . He began working and our communication was continued by writing. When he finished his MS thesis (1990), he asked me to let one of his friends to make his presentation, but I insisted that he has to do it only by pointing to the figures and tables (Fig.1). The examiners were satisfied (Fig. 2). It was a wonderful day . He was happy and thankful to all (Fig. 2). The same was done in the discussion of his Ph. D. (Fig 3), also on mycotoxins (1994). He continued to do research assending the academic ladder till he became a chief researcher and head of the Department of Mycology and Mycotoxins (2006-2009) and shared with me as co-supervisor on several theses and examiner of postgraduate students (Fig. 4) in the field of mycotoxins. Now he is the main researcher in this field in Veterinary Medicine in Egypt and he still works by himself in the laboratory (Fig. 5). I am happy that Prof. Atef Abdelaziz is co-author of this monograph M. Refai 3 Contents Introduction 5 Classification of mycotoxins 7 Mycotoxins and mycotoxigenic fungi 8 Aspegillus toxins 8 Keys for identification of Aspergillus groups 14 Common toxigenic aspergilli 16 Penicillium toxins 27 Description of the most common toxigenic Penicillium 29 Fusarium toxins 35 Description of some Fusarium species 37 Toxins of black fungi 40 Trichoderma toxins 45 Mycotoxicoses 46 Aspergillotoxicosis 46 Aflatoxicosis 46 Ochratoxicosis 49 Penicillotoxicosis 50 Fusariotoxicosis 51 Facial Eczema (Pithomycotoxicosis) 54 Stachybotryotoxicosis 55 Management of mycotoxins contamination in feed 56 Detection of aflatoxins 57 Prevention of mycotoxin formation58 Detoxification of mycotoxins in food and feedstuffs 60 Physical methods 60 Biological detoxification of mycotoxin 62 Control of mycotoxins during food processing 62 Limits for mycotoxins in foods and feeds 68 Future studies and recommendation 72 Synopsis on studies done by authors on mycotoxigenic fungi and mycotoxins in foods and feeds 73 References 159 Annexes 1171 Annex 1: Media , reagent and stains used for isolation , identification 171 Annex 2: Methods for Isolation, Enumeration and Identification of mycotoxigenic moulds 175 Annex 3 : Determination of mycotoxins in feed and food samples 178 Annex 4: Identification of fungal isolates by polymerase chain reaction (PCR):182 Annex 5: Rapid detection of mycotoxigenic fungi:187 4 1. Introduction Mycotoxins are secondary metabolites produced by filamentous fungi that have deleterious effects on human and animal consumers. They are structurally diverse, deriving from a number of biosynthetic pathways and their effect upon consumers is equally diverse ranging from acutely toxic to immunosuppressive or carcinogenic. The production of a particular mycotoxin is restricted to a limited number of fungal species and, in some instances, may be limited to particular strains within a species. Although over 400 mycotoxins have been described, relatively few are of major concern with respect to human and animal health, as they are responsible for the production of the great majority of the mycotoxins. The majority of these fungi infect plants and can be regarded as phytopathogens, Mycotoxin contamination is an economic problem for live stock and feed industries. The presence of mycotoxins in feedstuffs reduces the feed quality in terms of both energy and protein value.High moisture content (>12%) and grain damage favour mould growth. Mycotoxins may get concentrated from 30-500 times in broken grain as compared to whole grain. Risk from mycotoxin not only depends on the degree of contamination, but also on duration of exposure. There are no safe levels for mycotoxins as the damage depends on too many factors including health and nutritional status of the animal or bird, stress and other disease conditions and presence of other mycotoxins. Testing the feed for one particular toxin may not give a clear picture as there could be number of other mycotoxins. In practical feeding situations, it is rare to find a single mycotoxin. Mycotoxins generally have synergistic effects. So the damage caused by the combination is more devastating. Multiple mycotoxins at low to moderate levels may produce symptoms often distinctly different from those associated with individual mycotoxins. 5 Immunosuppression is one of the serious outcome of Mycotoxin contamination, often unnoticed, making animal or bird susceptible to infection and complex disease problems. Many mycotoxins are harmful to humans and animals when inhaled, ingested or brought into contact with human skin. Mycotoxins can cause a variety of short term as well as long-term health effects, ranging from immediate toxic response to potential long-term carcinogenic and teratogenic effects. Symptoms due to exposure to mycotoxins include dermatitis, cold and flu symptoms, sore throat, headache, fatigue, diarrhea, and impaired or altered immune function, which may lead to opportunistic infection. Mycotoxins are a known agent in biological warfare as a moderate illness compared to the other biologicals. 6 2. Classification of mycotoxins 2.1. Classification according to their mode of action: Cytotoxic: Patulin and penicillic acid. Emetic: Vomitoxin and T-2 toxin Immunosuppressive: Aflatoxin B1 Carcinogenic: Aflatoxins, fusarenon, patulin, rugulosin. Mutagenic: Aflatoxins, sterigmatocystin, trichothecenes. Oestrogenic: Zearalenon. Teratogenic: Aflatoxins, citrinin, ochratoxin A. 2.2. Classification according to the target organs: Hepatotoxins (liver tissues): Aflatoxins, ochratoxins, rubratoxins, sporidesmin, sterigmatocystin, luteoskyrin, rugulosin, islanditoxin. Nephrotoxins (kidney tissues): Citrinin, ochratoxin A, oxalic acid. Neurotoxins (nervous system tissues) : Citreoviridin, patulin, cyclopiazonic acid, fumitremorgen, territrems, verruculogen. Enterotoxins (alimentary organs and canal), dermotoxins(skin): Trichothecenes, verrucarin. 2.3. Classification according to the target systems: Vascular system :(Increased vascular fragility, hemorrhage into body tissues, or from lung, e. g., aflatoxin, satratoxin, roridins). Digestive system: (Diarrhea, vomiting, intestinal hemorrhage, liver effects, necrosis, fibrosis: aflatoxin; caustic effects on mucous membranes: T-2 toxin; anorexia: vomitoxin. 7 Respiratory system: Respiratory distress, bleeding from lungs e. g., trichothecenes. Nervous system, Tremors, incoordination, depression, headache, e. g.,tremorgens, trichothecenes. Cutaneous system: Rash, burning sensation sloughing of skin, photosensitization, e. g., trichothecenes. Urinary system: Nephrotoxicity, e. g. ochratoxin, citrinin. Reproductive system; Infertility, changes in reproductive cycles, e. g. T-2 toxin, zearalenone. Immune system: Changes or suppression: many mycotoxins. 3. Mycotoxins and mycotoxigenic fungi 3.1. Aspegillus

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