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1 Toxic Mold

2 Mold - Bible Leviticus 14: the priest is to go and examine it and, if the mold has spread in the house, it is a persistent defiling mold; the house is unclean. 45 It must be torn down--its stones, timbers and all the plaster--and taken out of the town to an unclean place

3 Fungi Eukaryotic organisms that have chitin (carbohydrate backbone) in cell wall: provides structural integrity Kingdom (fungi): yeast, molds and mushrooms Estimated 1.5 to 5 million species Most fungi grow as tubular filaments called hyphae (interwoven mass of hyphae is called mycelium) Reproduce and disperse by formation of spores Saprophytic fungi obtain their nutrients by secreting enzymes that decay plant materials Play a crucial role in nature by breaking down dead organisms and releasing their nutrients for reuse (e.g. into the soil)

4 Indoor Mold Growth Sequence of Events 1. Water intrusion / water damage 2. Substrate for mold growth: any cellulose (plant based) material: wood, paper, wallpaper, dry wall, ceiling tiles, carpet glue, dust, etc 3. Mold growth starts and continues as long as adequate moisture source 4. Production of spores and secondary metabolites: mycotoxins, volatile organic compounds, etc

5 Black Mold WDB

6 Black Mold (Stachybotrys) Ceiling

7 Fungi / Mold recovered from Water Damaged Buildings Anderson B et al, Appl Environ Microbiol 2011;77:

8 Detection of Mold DNA from Dust in WDB in New Orleans after Hurricane Katrina Bloom E et al, Indoor Air 2009;19:

9 Stachybotrys chartarum Generally considered the prototype of toxic black mold Found in water damaged buildings, not found outdoors Produces several macrocyclic trichothecene toxins: roridin E; satratoxins F, G and H; isosatratoxins F, G and F; verrucarins B and J among others Spores are large (4.5 microns) and have a slimy surface therefore are not readily airborne Readily produces fine fragments (small particulates) which are much smaller than the spores (< 2 microns) and easily become airborne Small fragments are released 500X higher concentration in the air than that of spores Both spores and fine fragments contain mycotoxins

10 Detection of Trichothecene Mycotoxins from Sera of Individuals Exposed to Stachybotrys chartarum in Indoor Environments Group 1 (n = 18) Documented exposure to Stachybotrys chartarum Numerous symptoms Trichothecenes present serum: 77.7% Group 2 (n = 26) Samples from individuals with exposure to unidentified molds Symptoms varied (were more minor) Trichothecenes present in serum: 36.6% Group 3 (n = 26) Individuals with no symptoms and no known mold / mycotoxin exposure Tricohthecenes present in serum: 3.8% Brasel TL et al, Arch Environ Health 2004;59:

11 Mold: Secondary Metabolites Not essential to maintaining life cycle of the mold Give the molds that produce them a competitive advantage over other molds and bacteria Higher organisms not targeted by mycotoxins but get caught in the crossfire Types of secondary metabolites: Mycotoxins Volatile organic compounds (VOCs) Extracellular enzymes Extracellular proteins

12 Mycotoxin-producing Fungi / Mold Species Mycotoxin Aspergillus flavus Aflatoxin (AT) A. parasiticus AT A. ochraceus Ochratoxin A (OTA) A. niger OTA A. carbonarius OTA A. versicolor Sterigmatocystin, gliotoxin A. fumigatus Gliotoxin Penicillium verrucosum OTA P. nordicum OTA P. chrysogenum OTA Penicillium spp (many) Patulin, tremorgens Stachybotrys chartarum Macrocyclic trichothecenes (MT) Trichoderma viridae MT Chaetomium globosum Chaetoglobsin A, C

13 Examples of Common Mycotoxins Aflatoxin B1, Ochratoxin A and a Macrocyclic Trichothecene

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15 Mycotoxins Mechanisms of toxicity Bind to DNA and RNA Alters protein synthesis and functions (e.g. enzymes) Oxidative stress (e.g. lipid peroxidation) Depletes antioxidants Alters cell membrane function and transport Potent mitochondrial toxins (numerous mechanisms) Alters apoptosis

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17 Detection of Mycotoxins in CFS Patients

18 Other Diagnoses in Mycotoxin Cases Brewer, et al. Toxins 2013;5:

19 Detection of Mycotoxins in CFS Patients Brewer, et al. Toxins 2013;5:

20 Detection of Mycotoxins in CFS Patients Compared to Healthy Controls Brewer, et al. Toxins 2013;5:

21 Chronic Fatigue Syndrome Basic Science and Clinical Associations Immune dysregulation: abnormal cytokine release, inflammation, autoimmune features, immune deficiency Neurologic and neurocognitive abnormalities: CNS, peripheral nervous system and autonomic nervous system Endocrine abnormalities: especially HPA axis, but also thyroid, sex hormones Psychiatric / behavioral Oxidative stress: increased free radicals and impaired antioxidant function Mitochondrial dysfunction: shown in vitro and in clinical studies

22 Chronic Fatigue Syndrome Why consider mycotoxins? Clinical features (symptoms) of mycotoxin exposure and CFS similar Sick building syndrome has led to CFS (Chester and Levine, 1994) Mycotoxins are associated with numerous neurologic abnormalities Mycotoxins can lead to endocrine abnormalities Mycotoxins can lead to immune dysregulation Mycotoxins can lead to oxidative stress, free radicals and impaired antioxidant function Mycotoxins induce mitochondrial dysfunction

23 CFS and Mitochondria Mitochondrial abnormalities in CFS patients Impaired oxidative phosphorylation Reduced ATP stores Impaired recycling of ADP to ATP Increased lactic acid with exercise Reduced oxidative muscle metabolism Mitochondrial DNA deletions Mitochondrial degeneration on muscle biopsy Increased generation of free radicals Diminished carnitine levels

24 Mycotoxins and Mitochondria Aflatoxin: mitochondrial DNA adducts, inhibit protein synthesis, pleomorphism, disruption of cristae, membrane damage, induction of apoptosis Trichothecenes: oxidative stress, apoptosis, inhibition of protein, RNA and DNA synthesis, opening of PtCP, loss of transmembrane potential and mitochondrial translation Ochratoxin A: membrane swelling, disarray of cristae, loss of transmembrane potential, inhibition of succinate cytochrome c reductase and dehydrogenase, inhibition of succinate-supported electron transfer, inhibition of activities of the respiratory chain, oxidative stress, DNA damage, apoptosis

25 Mycotoxin-related Chronic Illness Input / Output Model Input of mycotoxins into the body External: ongoing exposure or re-exposures in buildings the patient lives, works or both Internal: mold internally releases mycotoxins Output of mycotoxins out of the body Kidneys urine Liver / Bile fecal (issue of reabsorption and enterohepatic recirculation) Sweat

26 Mycotoxin-related Chronic Illness Potential Treatments Reduce / eliminate input Eliminate ongoing exposure or re-exposure from environment (living environment, work or both) Antifungal drugs to eliminate internal mold Enhance output Mycotoxin binders Increase sweating Increase detoxification clearance (e.g. glutathione)

27 Mycotoxin-related Chronic Illness Antifungals Triazoles: itraconazole, voriconazole, posaconazole Ecchinocandins: micafungin, caspofungin Polyenes: amphotericin B (topical)

28 Mycotoxin-related Chronic Illness Mycotoxin binders Cholestyramine Activated charcoal Others: chitosan, bentonite, glucomannan, etc

29 Mycotoxin-related Chronic Illness Case history of remote exposure: Exposure in damp / moldy home over 20 years ago Symptoms started while living in that home Hasn t lived in the original home or been in an environment of concern for mold exposure for at least 15 years Has had chronic symptoms since the original home (over 20 years) Random urine specimen sent to RTL > 20 years after onset of symptoms: demonstrates elevated levels of mycotoxins

30 Mycotoxin-related Chronic Illness Hypothesis: There are mycotoxin producing fungi / molds internally that are continuously releasing mycotoxins Questions: Where are the internal molds? Answer: paranasal sinuses (mycotoxin-associated fungal organisms in biofilm) How can the internal molds be eradicated? Answer: direct treatment of the sinuses with intranasal therapy (+/- systemic antifungal agents)

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32 Mycotoxin-related Chronic Illness Why consider the sinuses as chronic reservoir? Fungi are almost always present in the sinuses of healthy population and chronic rhinosinusitis (CRS) cases Mycotoxin producing fungal species have been found in the sinuses Mycotoxins can be found in the urine of CRS cases Mycotoxins can be recovered from nasal washings in mold exposure cases, as well as sinus tissue specimens Fungi in the sinuses readily form biofilm (bacterial component may be involved) Mycotoxin producing fungi in the sinuses are likely embedded in biofilm Direct treatment of the sinuses (intranasal antifungals) improve symptoms

33 Nasa Touch

34 Intranasal Therapy Antifungal: amphotericin B, itraconazole Biofilm buster : chelating agent (e.g. EDTA) Corticosteroids: reduce inflammation if there is history of chronic rhinosinusitis and / or allergies Antibiotics: may be helpful to reduce bacterial component (example: mupirocin)

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