Inhibition of dopamine conversion to norepinephrine by Clostridia metabolites appears to be a (the) major cause of autism, schizophrenia, and other neuropsychiatric disorders.

William Shaw, PhD

Concentrations of the dopamine metabolite homovanillic acid, or HVA, have been reported to be much higher in the urine of children with autism compared to controls. In the same study, severity of autism symptoms was directly related to the concentration of HVA. There was a relation between the urinary HVA concentration and increased agitation, stereotypical behaviors, and reduced spontaneous behavior. Furthermore, vitamin B6, which has been shown to decrease autistic symptoms, decreases urinary HVA concentrations. Excess dopamine has been implicated in the etiology of psychotic behavior and schizophrenia for over 40 years. Drugs that inhibit dopamine binding to dopaminergic receptors have been some of the most widely used pharmaceuticals used as antipsychotic drugs and have been widely used in the treatment of autism. Recent evidence reviewed below indicates that dopamine in high concentrations may be toxic to the brain.

Dopamine is a very reactive molecule compared with other neurotransmitters, and dopamine degradation naturally produces oxidative species (Figure 1). More than 90 percent of dopamine in dopaminergic neurons is stored in abundant terminal vesicles and is protected from degradation. However, a small fraction of dopamine is cytosolic, and it is the major source of dopamine metabolism and presumed toxicity. Cytosolic dopamine (Figure 1) undergoes degradation to form 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) via the monoamine oxidase pathway. Alternatively, dopamine undergoes oxidation in the presence of excess iron or copper (common in autism and schizophrenia) to form dopamine cyclized o-quinone, which is then converted to dopamine cyclized o-semiquinone, depleting NADPH in the process. Dopamine cyclized o-semiquinone then reacts with molecular oxygen to form oxygen superoxide free radical, an extremely toxic oxidizing agent. In the process, dopamine cyclized o-quinone is reformed, resulting in a vicious cycle extremely toxic to tissues producing dopamine, including the brain, peripheral nerves, and the adrenal gland.

 It is estimated that each molecule of dopamine cyclized o-quinone produces thousands of molecules of oxygen superoxide free radical in addition to depleting NADPH. The o-quinone also reacts with cysteine residues on glutathione or proteins to form cysteinyl-dopamine conjugates (Figure 1). One of these dopamine conjugates is converted to N-acetylcysteinyl dopamine thioether, which causes apoptosis (programmed cell death) of dopaminergic cells. These biochemical abnormalities cause severe neurodegeneration in pathways that utilize dopamine as a neurotransmitter. Neurodegeneration is due to depletion of brain glutathione and NADPH as well as the overproduction of oxygen superoxide free radicals and neurotoxic N-acetylcysteinyl dopamine thioether. In addition, the depletion of NADPH also results in a diminished ability to convert oxidized glutathione back to its reduced form.

What is the likely cause of elevated dopamine in autism? A significant number of studies have documented increased incidence of stool cultures positive for certain species of Clostridia bacteria in the intestine in children with autism using culture and PCR techniques. All these studies have indicated a disproportionate increase in various Clostridia species in stool samples compared to normal controls. In addition, metabolic testing has identified the metabolites 3-(3-hydroxyphenl)-3-hydroxypropionic acid (HPHPA) and 4-cresol from Clostridia bacteria at significantly higher concentrations in the urine samples of children with autism and in schizophrenia.

Treatment with antibiotics against Clostridia species, such as metronidazole and vancomycin, eliminates these urinary metabolites with reported concomitant improvement in autistic symptoms. In addition, I had noticed a correlation between elevated HPHPA and elevated urine homovanillic acid (HVA). The probable mechanism for this correlation is that certain Clostridia metabolites have the ability to inactivate dopamine beta-hydroxylase, which is needed for the conversion of dopamine to norepinephrine (Figure 2).

Figure 2. Effect of Clostridia metabolites on human catecholamine metabolism. DHPPA, 4-cresol, HPHPA, HVA, and VMA are all measured in The Great Plains Laboratory organic acid test.

Such metabolites are not found at only trace levels. The concentration of the Clostridia metabolite HPHPA in children with autism may sometimes exceed the urinary concentration of the norepinephrine metabolite vanillylmandelic acid (VMA) by a thousand fold on a molar basis and may be the major organic acid in urine in those with severe gastrointestinal Clostridia overgrowth, and even exceed the concentration of all the other organic acids combined. Dopamine beta hydroxylase that converts dopamine to norepinephrine in serum of severely intellectually disabled children with autism was much lower than in those who were higher functioning. Decreased urine output of the major norepinephrine metabolite meta-hydroxyphenolglycol (MHPG) was decreased in urine samples of children with autism, consistent with inhibition of dopamine beta hydroxylase.

Many physicians treating children with autism have noted that the severity of autistic symptoms is related to the concentration of the Clostridia marker 3-(3-hydroxyphenyl)-3-hydroxypropionic acid (HPHPA) in urine. These are probably the children with autism with severe and even psychotic behavior treated with Risperdal® and other anti-psychotic drugs, which block the activation of dopamine receptors by excess dopamine. I have identified a number of species of Clostridia species that produce HPHPA including C. sporogenes, C.botulinum, C. caloritolerans, C. mangenoti, C. ghoni, C.bifermentans, C. difficile, and C. sordellii. All species of Clostridia are spore formers and thus may persist for long periods of time in the gastrointestinal tracts even after antibiotic treatment with oral vancomycin and metronidazole.

How do the changes in brain neurotransmitters caused by Clostridia metabolites alter behavior? The increase in phenolic Clostridia metabolites common in autism significantly decreases brain dopamine beta hydroxylase activity. This leads to overproduction of brain dopamine and reduced concentrations of brain norepinephrine, and can cause obsessive, compulsive, stereotypical behaviors associated with brain dopamine excess and reduced exploratory behavior and learning in novel environments that are associated with brain norepinephrine deficiency. Such increases in dopamine in autism have been verified by finding marked increases in the major dopamine metabolite homovanillic acid (HVA) in urine. The increased concentrations of HVA in urine samples of children with autism are directly related to the degree of abnormal behavior. The concentrations of HVA in the urine of some children with autism are markedly abnormal.

In addition to alteration of brain neurotransmitters, the inhibition of the production of norepinephrine and epinephrine by Clostridia metabolites may have a prominent effect on the production of neurotransmitters by the sympathetic nervous system and the adrenal gland. The major neurotransmitter of the sympathetic nervous system that regulates the eyes, sweat glands, blood vessels, heart, lungs, stomach, and intestine is norepinephrine. An inadequate supply of norepinephrine or a substitution of dopamine for norepinephrine might result in profound systemic effects on physiology. The adrenal gland which produces both norepinephrine and epinephrine might also begin to release dopamine instead, causing profound alteration in all physiological functions. In addition to abnormal physiology caused by dopamine substitution for norepinephrine and dopamine, dopamine excess causes free radical damage to the tissues producing it, perhaps leading to permanent damage of the brain, adrenal glands, and sympathetic nervous system if the Clostridia metabolites persist for prolonged periods of time, if glutathione is severely depleted, and if there is apoptotic damage caused by the dopamine metabolite N-acetylcysteinyl dopamine thioether.

Depletion of glutathione can be monitored in The Great Plains Laboratory organic acid test by tracking the metabolite pyroglutamic acid, which is increased in both blood and urine when glutathione is depleted. In addition, The Great Plains Laboratory also tests the other molecules involved in this toxic pathway, the dopamine metabolite homovanillic acid (HVA), the epinephrine and norepinephrine metabolite VMA and the Clostridia metabolites HPHPA and 4-cresol.

In summary, gastrointestinal Clostridia bacteria have the ability to markedly alter behavior in autism and other neuropsychiatric diseases by production of phenolic compounds that dramatically alter the balance of both dopamine and norepinephrine. Excess dopamine not only causes abnormal behavior but also depletes the brain of glutathione and NADPH and causes a vicious cycle producing large quantities of oxygen superoxide that causes severe brain damage. Such alterations appear to be a (the) major factor in the causation of autism and schizophrenia. The organic acid test (see sample organic acid test report below) now has the ability to unravel a major mystery in the causation of autism, schizophrenia, and other neuropsychiatric diseases, namely the reason for dopamine excess in these disorders.

In the past, some physicians would order the organic acid test once a year or less. With the new knowledge of the mechanism of Clostridia toxicity via inhibition of dopamine beta-hydroxylase, it seems that the control of such toxic organisms needs to monitored much more frequently to prevent serious brain, adrenal gland, and sympathetic nervous system damage caused by excess dopamine and oxygen superoxide. Below is a test report of a child with autism tested with The Great Plains Laboratory Organic acid test.

DISCUSSION OF PATIENT RESULTS
In the graph above, the vertical bar is the upper limit of normal and the patient’s value is plotted inside a diamond (red for abnormal, black for normal). The above results were from a boy with severe autism. The HPHPA Clostridia marker was very high (979 mmol/mol creatinine), about 4.5 times the upper limit of normal. However, the metabolite due to Clostridium difficile was in the normal range, indicating that Clostridium difficile was unlikely to be the Clostridium bacteria producing the high HPHPA. In other words, a different Clostridia species was implicated. The major dopamine metabolite homovanillic acid (HVA) was extremely high (87 mmol/mol creatinine), almost 7 times the upper limit of normal. The major metabolite of epinephrine and norepinephrine, VMA was in the normal range. The HVA/VMA ratio was 15, more than five times higher than the upper limit of normal, indicating a severe imbalance in the production of epinephrine/norepinephrine and that of dopamine. The very high dopamine metabolite, HVA, indicates that the brain, adrenal glands, and sympathetic nervous system may be subject to severe oxidative stress due to superoxide free radicals and that brain damage due to severe oxidative stress might result if the Clostridia bacteria are left untreated. Below the same patient’s results are displayed in a form that is related to the metabolic pathways. This graphical result now appears on all organic acid results from The Great Plains Laboratory, Inc.

Reference:

1. Shaw W. Increased urinary excretion of a 3-(3-hydroxyphenyl)-3-hydroxypropionic acid (HPHPA), an abnormal phenylalanine metabolite of Clostridia spp. in the gastrointestinal tract, in urine samples from patients with autism and schizophrenia. Nutr Neurosci. 2010 Jun;13(3):135-43.

The Clinical Significance of Organic Acids Testing to Mental Health – How Fungal, Bacterial, Mitochondrial, and Other Test Markers Influence the Brain

Kurt Woeller, D.O.

Organic acids testing is diagnostic tool that every healthcare practitioner should know about.  Whether you are a family practitioner, psychiatrist, a nutritionist, or other type of practitioner, the information provided by organic acids testing can help identify underlying causes of a variety of chronic illnesses, including the symptoms of autism, neuropsychiatric disorders like depression and anxiety, and neurodegenerative disorders like Alzheimer’s disease.  Below is a review of some of the most clinically significant markers measured with organic acids testing to mental health and the health of the brain in general. 

Many of the case studies reviewed in presentations about organic acids testing involve patients with autism.  While autism may not typically be considered a mental health disorder, it is a neurodevelopmental disorder and many autistic individuals suffer with mental symptoms such as anxiety and depression, along with associated behavioral problems.  Many patients with autism also have mitochondrial dysfunction and chronic infections (like Candida and clostridia), which are measured with organic acids testing. (1)

Mitochondria are linked to every organ system in the body, including the brain, and there markers for mitochondrial function in organic acids testing. Without adequate mitochondrial function, neurons cannot function appropriately to produce neurochemicals such as dopamine and serotonin.  Mitochondria are damaged by various endogenous toxins produced by Candida (a fungus) such as tartaric acid and citramalic acid. Also, certain clostridia bacteria produce propionic acid which damages mitochondria. Candida and clostridia are both measured with organic acids testing.  Mitochondria are also damaged by oxalate, which is produced by Candida and some molds, and is also measured with organic acids testing. Certain molds like Aspergillus produce mycotoxins which directly damage mitochondria. Organic acids testing specifically measures candida toxins, bacteria toxins, and mold toxins, along with mitochondria markers. (2, 3)

Clostridia bacteria can produce various compounds like HPHPA, 4-Hydroxyphenylacetic acid and 4-Cresol (all measured with organic acids testing), and are known to inhibit dopamine metabolism. These chemicals inhibit Dopamine-Beta Hydroxylase which causes neuronal dopamine levels to rise. This has been associated with paranoia and schizophrenia. Also, the breakdown products of dopamine are neurotoxic and cause brain receptor damage.  Chronic infections and the compounds produced from them such as bacteria lipopolysaccharides (LPS), along with elevated cortisol (seen in hypothalamic-pituitary-adrenal dysfunction), viral infections, and beta-amyloid and niacin deficiency (seen in schizophrenia) can trigger tryptophan metabolism problems. Tryptophan is the amino acid precursor to serotonin. In the presence of these chronic stressors, tryptophan conversion to serotonin is reduced. This can lead to depression and anxiety. Elevated tryptophan metabolites can lead to increased quinolinic acid (QA).  (4)

Quinolinic acid is neurotoxic and measured with organic acids testing. It is an NDMA receptor agonist, which is linked to various mental health disorders (anxiety, depression, suicidal ideation) and chronic neurodegenerative diseases (Alzheimer’s, Huntington’s). Quinolinic acid can also block acetylcholine production (linked to memory) and gamma-amino-butyric acid (which can trigger anxiety and panic). (5)

The aforementioned markers in organic acids testing are some of the most clinically significant to mental health and brain function, though there are many other examples.  This information is critical for mental health professionals to help deepen their knowledge about sophisticated testing and advanced solutions for patient intervention. 

References

1. Shaw, W., et. al. Increased Urinary Excretion of Analogs of Krebs Cycle Metabolites and Arabinose in Two Brothers with Autistic Features. Clin Chem 41:1094-1104, 1995.

2. Shaw, W., et. al. Assessment of antifungal drug therapy in autism by measurement of suspected microbial metabolites in urine with GC/MS. Clinical Practice of Alternative Medicine: 15-26.

3. Persico AM, et. al. Urinary p-cresol in autism spectrum disorders, Neurotoxicol Teratol. 2013 Mar-Apr;36:82-90, 2012 Sep 10.

4. Heyes MP, et. al. A mechanism of quinolinic acid formation by brain in inflammatory neurological disease. Attenuation of synthesis from L-tryptophan by 6-chlorotryptophan and 4-chloro-3-hydroxyanthranilate. Brain. 1993 Dec;116 (pt 6):1425-50.

5. Ganiyu Oboh, et. al. Anticholinesterase and Antioxidative Properties of Aqueous Extract of Cola acuminata Seed In Vitro. Int J Alzheimers Dis. 2014; 2014: 498629.

Magnesium: The Missing Link in Mental Health?

by James Greenblatt, MD

Chief Medical Officer at Walden Behavioral Care in Waltham, MD
Assistant Clinical Professor of Psychiatry at Tufts University School of Medicine and Dartmouth College Geisel School of Medicine

Magnesium is a cofactor in more than 325 enzymatic reactions—in DNA and neurotransmitters; in the bones, heart and brain; in every cell of the body. Unfortunately, a deficiency of this crucial mineral is the most common nutritional deficiency I see in my practice as an integrative psychiatrist. Fortunately, supplementation with magnesium is the most impactful integrative treatment I use, particularly in depression and attention deficit hyperactivity disorder (ADHD).

Why is magnesium deficiency so common, and why is restoring the mineral so essential to mental and emotional well-being and behavioral balance? The rest of this article addresses those two questions, and presents aspects of my therapeutic approach.

Magnesium Deficiency

 The population is deficient in magnesium—found abundantly in whole grains, beans and legumes, nuts and seeds, and leafy greens, as well as cocoa and molasses—for several reasons.

Soil depletion. Intensive agricultural practices rob the soil of magnesium and don’t replace it. As a result, many core food crops—such as whole grains—are low in magnesium. A recent paper in Crop Journal put it this way: Magnesium’s “importance as a macronutrient ion has been overlooked in recent decades by botanists and agriculturists, who did not regard Mg deficiency in plants as a severe health problem. However, recent studies have shown, surprisingly, that Mg contents in historical cereal seeds have markedly declined over time, and two thirds of people surveyed in developed countries received less than their minimum daily Mg requirement.” [1]  

Food processing. Magnesium is stripped from foods during food processing. For example, refined grains—without magnesium-rich germ and bran—have only 16% of the magnesium of whole grains. [2]

Stress. Physical and emotional stress—a constant reality in our 24/7 society—drain the body of magnesium. In fact, studies show inverse relationships between serum cortisol and magnesium—the higher the magnesium, the lower the cortisol. Stress robs the body of magnesium—but the body must have magnesium to respond effectively to stress.

Other factors. Many medications—such as medications for ADHD—deplete magnesium. So does the intake of alcohol, caffeine and soft drinks.

The result: In 1900, the average intake of magnesium was 475 to 500 mg daily. Today, it’s 175 to 225 mg daily. Which means that only one-third of adult Americans get the daily RDA for magnesium—320 mg for women, and 420 mg for men. (And many researchers consider the RDA itself inadequate.)  And that magnesium deficit causes deficits in health. Magnesium deficiency has been cited as contributing to atherosclerosis, hypertension, type 2 diabetes, obesity, osteoporosis and certain types of cancer. [4] But detecting that deficiency in laboratory testing is difficult, because most magnesium in the body is stored in the skeletal and other tissues. Only 1% is in the blood, so plasma levels are not a reliable indicator. That means a “normal” magnesium blood level may exist despite a serious magnesium deficit. An effective therapeutic strategy: Assume a deficit is present, and prescribe the mineral along with other appropriate medical and natural treatments. That’s particularly true if the patient has symptoms such as anxiety, irritability, insomnia and constipation, all of which indicate a magnesium deficiency.

The Mind Mineral

Some of the highest levels of magnesium in the body are found in the central nervous system, with studies dating back to the 1920s showing how crucial magnesium is for a balanced brain…

It’s known, for example, that magnesium interacts with GABA receptors, supporting the calming actions of this neurotransmitter. Magnesium also keeps glutamate—an excitatory neurotransmitter—within healthy limits. Patients with higher magnesium levels also have healthy amounts of serotonin in the cerebrospinal fluid. And the synthesis of dopamine requires magnesium.

In summary, the body needs magnesium to create neurotransmitters (biosynthesis) and for those neurotransmitters to actually transmit. Magnesium also acts at both the pituitary and adrenal levels. In the pituitary gland, it modulates the release of ACTH, a hormone that travels to the adrenal glands, stimulating cortisol release. In the adrenal gland, it maintains a healthy response to ACTH, keeping cortisol release within a normal range. As a result, magnesium is a must for maintaining the homeostasis of the HPA axis. Given all these key mechanisms of action, it’s not surprising that a lack of the mineral can produce psychiatric and other types of problems. The patient may have: Difficulty with memory and concentration. Depression, apathy and fatigue. Emotional lability. Irritability, nervousness and anxiety. Insomnia. Migraine headaches. Constipation. PMS. Dysmenorrhea. Fibromyalgia. Autism. ADHD. Fortunately, studies show that magnesium repletion—restoring normal levels of the mineral—produces positive changes in mood and cognition, healthy eating behavior, healthy stress responses, better quality of sleep, and better efficacy of other modalities, such as medications. Let’s look at two areas in which magnesium supplementation is particularly effective: Depression and ADHD.

Depression

A cross-sectional, population-based data set—the National Health and Nutrition Examination Survey—was used to explore the relationship of magnesium intake and depression in nearly 9,000 US adults. Researchers found significant association between very low magnesium intake and depression, especially in younger adults. [5] And in a recent meta-analysis of 11 studies on magnesium and depression, people with the lowest intake of magnesium were 81% more likely to be depressed than those with the highest intake. [6] In a clinical study of 23 senior citizens with depression, low blood levels of magnesium and type 2 diabetes, magnesium was compared to the standard antidepressant medication imipramine (Tofranil)—one group received 450 mg of magnesium daily and one group received 50 mg of imipramine. After 12 weeks, depression ratings were equally improved in both groups. [7] In my practice, I nearly always prescribe magnesium to a patient with diagnosed depression. You can read more about the integrative approach to depression in Integrative Therapies for Depression: Redefining Models for Assessment, Treatment and Prevention (CRC Press), which I co-edited, and in Breakthrough Depression Solution: Mastering Your Mood with Nutrition, Diet & Supplementation (Sunrise River Press, 2nd Edition).

Attention Deficit Hyperactivity Disorder

Magnesium deficiency afflicts 90% of all people with ADHD and triggers symptoms like restlessness, poor focus, irritability, sleep problems, and anxiety. These symptoms can lessen or vanish one month after supplementation starts. Magne­sium can also prevent or reverse ADHD drug side effects. That’s why all of my ADHD patients get a prescription for magnesium. For adolescents, I typically prescribe 200 mg, twice daily. For children 10 to 12, 100 mg, twice daily. For children 6 to 9, 50 mg, twice daily. Typically, I recommend magnesium glycinate, using a powdered product. I describe my entire approach to magnesium and ADHD (and to the disorder’s overall integrative treatment) in my book Finally Focused: The Breakthrough Natural Treatment Plan for ADHD That Restores Attention, Minimizes Hyperactivity, and Helps Eliminate Drug Side Effects. (Forthcoming from Harmony Books in May 2017)

Dosage and Form

I have found that 125 to 300 mg of magnesium glycinate at meals and a bedtime (four times daily) produces clinically significant benefits in mood. (This form of magnesium is gentle on the digestive tract.) 200 to 300 mg of magnesium glycinate or citrate before bed supports sleep onset and duration through the night. You can also find magnesium in powder or liquid form, which are effective alternatives to capsules, particularly for children with ADHD. Ways to increase the bioavailability of magnesium include: Supplementing with vitamin D3, which increases cellular uptake of the mineral. Vitamin B6 also helps magnesium accumulate in cells. Taking the mineral in divided doses instead of a single daily dose. Taking it with carbohydrates, with improves absorption from the intestine. And taking an organic form, such as glycinate or citrate, which improves absorption by protecting the mineral from antagonists in the digestive tract. Avoid giving magnesium in enteric-coated capsules, which decreases absorption in the intestine.

Magnesium oxide is poorly absorbed and tends to cause loose stools. Magnesium-l-threonate has been shown to readily cross the blood-brain barrier, and animal studies show that it supports learning ability, short and long-term memory and brain function, I don’t typically prescribe it, however, because of its higher cost, and the clinical effectiveness of other forms. The therapeutic response to magnesium typically takes several weeks, as levels gradually increase in the body.

CITATIONS

[1] Guo W., et al. Magnesium deficiency in plants: An urgent problem. The Crop Journal, Volume 4, Issue 2, April 2016, Pages 83-91.

[2] http://www.ancient-minerals.com/magnesium-sources/dietary/

[3] https://www.washingtonpost.com/national/health-science/magnesium-is-essential-to-your-health-but-many-people-dont-get-enough-of-it/2017/06/09/77bc35b4-2515-11e7-bb9d-8cd6118e1409_story.html?noredirect=on&utm_term=.b92d507bf92a

[4] Volpe, SL. Magnesium in Disease Prevention and Overall Health. Advances in Nutrition, 2013 May; 4(3): 378S-383S.

[5] Tarleton EK, at al. Magnesium Intake in Depression in Adults. Journal of the American Board of Family Medicine, 2015 Mar-Apr;28(2):249-56.

[6] Li B, et al. Dietary magnesium and calcium intake and risk of depression in the general population: A meta-analysis. Australian and New Zealand Journal of Psychiatry, 2016 Nov 1. [Epub ahead of print].

[7] Barragan-Rodriquez L, et al. Efficacy and safety or oral magnesium supplementation in the treatment of depression in the elderly with type 2 diabetes: a randomized, equivalent trial. Magnesium Research, 2008 Dec;21(4):218-23.

Clostridia Detection and Comparison of Organic Acid Detection Versus Stool Testing

William Shaw, Ph.D.

Continued research at The Great Plains Laboratory has resulted in new information on Clostridia bacteria markers that will soon be available for the urine organic acid test. New information will soon be available for the organic acid interpretations of 3 (3 hydroxyphenyl)-3 hydroxypropionic acid (HPHPA), 4-hydroxyphenylacetic acid, phenyllactic acid, and 3-indoleacetic acid at the beginning of 2015.

In addition, this article will help to clarify information about the increased value of organic acid testing compared to stool testing for assessing Clostridia species.

HPHPA

First, the species that are the major producers of the precursors of HPHPA have been identified and include C. botulinum, C. sporogenes, and C.caloritolerans. (It is common to use the abbreviation for the Clostridia genus "C" when giving the genus and species designation.)

C. botulinum is a gram-positive, rod-shaped, anaerobic, spore-forming, motile bacterium with the ability to produce the neurotoxin botulinum. The botulinum toxin can cause a severe flaccid paralytic disease in humans and animals and is the most potent toxin known to humankind (natural or synthetic) with a lethal dose of less than 1 μg (microgram) in humans. Symptoms of botulism include weakness, trouble seeing, feeling tired, and trouble speaking. This may then be followed by weakness of the arms, chest muscles, and legs. In food borne botulism, symptoms generally begin 18 to 36 hours after eating a contaminated food, but they can occur as early as 6 hours or as late as 10 days after eating the food.

It is interesting that the symptoms of botulism vary widely from a mild illness for which the patient may seek no medical treatment to a fulminant disease, killing within 24 hours (1). Since laboratory testing for this organism is only available at state health departments, it seems likely that many cases of botulism, especially the mild cases, may be undiagnosed. I suspect that some children with autistic behavior,with extremely high urine HPHPA, little or no speech, and extremely severe low muscle tone might actually have undiagnosed botulism, and further research on this possibility is warranted.

C. sporogenes is virtually identical to C. botulinum except it is lacking the gene for the botulinum neurotoxin. Like C. botulinum, it is an anaerobic gram-positive, rod-shaped bacterium that produces oval, subterminal endospores, and is commonly found in soil.

C. caloritolerans is named after its extreme heat (calor) resistance (tolerans). It can survive at the boiling point for 8 hours (2); its ability to resist heat may allow transmission even in well-cooked food. No scientific papers on any disease associations (other than my own articles dealing with its production of HPHPA) were found, which means there is still a great deal of research opportunity for microbiologists in the future.

4-Hydroxyphenylacetic Acid

High 4-hydroxyphenylacetic acid is associated with small intestinal bacteria overgrowth due to its production by the following Clostridia bacteria: C. diificile, C. stricklandii, C. lituseburense, C. subterminale, C. putrefaciens, and C. propionicum. C. difficile can be distinguished from the other species by its production of 4-cresol; none of the other species produce 4-cresol. No information on the pathogenicity of the other species producing 4-hydroxyphenylacetic acid is available. However, it is likely that 4-hydroxyphenylacetic is also an inhibitor of dopamine-beta-hydroxylase and appropriate treatment with probiotics or antibiotics may be clinically useful. 4-hydroxyphenylacetic acid is associated with bacterial overgrowth of the small intestine (3). Elevated values are common in celiac disease and cystic fibrosis, and have also been reported in jejuna web, transient lactose intolerance, Giardia infection, ileal resection, ileo-colic intersusseception, septicemia, and projectile vomiting. The elevations of 4-hydroxyphenylacetic acid in celiac disease and cystic fibrosis are so prevalent that involvement of these Clostridia bacteria may play a role in these illnesses. In C. difficileinfections 4-hydroxyphenylacetic acid is utilized by this bacteria to produce 4-cresol.

Phenyllactic Acid

Very high amounts of phenyllactic acid are found in the rare genetic disease phenylketonuria (PKU). Moderate amounts of phenyllactic acid may be due to gastrointestinal overgrowth of the intestine of the following Clostridia bacteria: C. sordellii, C. stricklandii, C. mangenoti, C. ghoni, and C. bifermentans. C sordellii is usually considered a nonpathogen except in immunocompromised people, but has been implicated in catastrophic infectious gynecologic illnesses among women of childbearing age. The other species have rarely or never been reported to be pathogenic.

3-Indoleacetic Acid

High 3-indoleacetic acid in urine is a byproduct of C. stricklandii, C. lituseburense, C. subterminale, and C. putrefaciens. No information on the pathogenicity of these species producing indoleacetic acid is available. However, very high amounts of this metabolite derived from tryptophan might indicate a depletion of tryptophan needed for other physiological functions.

4-Cresol

4-cresol is predominantly produced by C. difficile, a pathogenic bacteria, that is one of the most common pathogens spread in hospitals. Toxin-producing strains of C. difficile can cause illness ranging from mild or moderate diarrhea to pseudomembranous colitis, which can lead to toxic dilatation of the colon (megacolon), sepsis, and death (4). 4-cresol (para-cresol) has been used as a specific marker for Clostridium difficile (5). 4-Cresol, a phenolic compound, is classified as a type-B toxic agent and can cause rapid circulatory collapse and death in humans (6). Yokoyama et al. (7) have recently proposed that intestinal production of 4-cresol may be responsible for a growth-depressing effect on animals. Signs of acute toxicity in animals typically include hypoactivity, salivation, tremors and convulsions. High amounts of 4-cresol have been found in autism (8); the amount of 4- cresol in the urine has been found elevated in baseline samples and in replica samples of autistic children. Higher values of 4-cresol are found in girls with autism compared to boys with autism and higher values are associated with greater clinical severity of autistic symptoms and history of behavioral regression. 4-cresol is apparently produced by Clostridia difficile as an antimicrobial compound that kills other species of bacteria in the gastrointestinal tract, allowing the Clostridia difficile to proliferate and predominate.

Organic acid test superior to stool testing for Clostridia testing

C. difficile is the only species of 100 species of Clostridia from the gastrointestinal tract to be commonly tested in hospital laboratories throughout the world. However, this species is not commonly cultured, but rather is detected by its toxin formation. The gastrointestinal damage caused by C. difficile is thought to be due to exposure to two toxins produced by C. difficile, toxin A and toxin B, with toxin B considered to be more toxic (4). The toxins can be tested by immunoassay of stool samples which is a fairly rapid test. Toxigenic stool culture, which requires growing the bacteria in a culture and detecting the presence of the toxins, is the most sensitive test for C. difficile, and it is still considered to be the gold standard (4). However, it can take 2 to 3 days for results. Polymerase chain reaction (PCR) evaluation of the C. difficile toxins is also becoming more available. Virtually all of the research on C. difficile is related to the effects of this species of bacteria on the intestinal tract. Toxin-negative C. difficile strains are considered nonpathogenic for the infection of the intestine (4) but cresol producing strains that don't produce toxins and B may be pathogenic due to their effects on brain metabolism and for the inherent toxicity of 4-cresol itself.

In addition, urinary 4-cresol elevations associated with C. difficileovergrowth are much less common than urinary HPHPA elevations associated with other Clostridia species. In a survey of 1000 consecutive samples submitted for urine organic acids tests, The Great Plains Laboratory found that 15.2% were abnormally elevated for HPHPA, 6.8% were abnormally elevated for 4-cresol, and 1.6% were abnormally elevated for both HPHPA and 4-cresol for a total positive percentage of 23.6%. Thus, if only stool testing for Clostridium difficile is performed on a patient, at least 15.2/23.6 or 64.4% (nearly two-thirds) of patients with clinically significant infections with other types of Clostridia might be missed.

Sometimes total Clostridia are tested using culture methods or PCR (polymerase chain reaction) technology. In one case, a parent showed me the stool test results of their child with autism. They had done a stool test with a laboratory using PCR technology to determine both C. difficile and total Clostridia. The total Clostridia was reported as extremely low and the C. difficile negative, but The Great Plains Laboratory organic acid test found high levels of the HPHPA marker. If the parent had relied on the stool test alone, their child might have missed an important therapeutic intervention that can restore normal neurotransmitter balance. The advantage of The Great Plains Laboratory organic acid test is that it is not necessary to determine particular species of Clostridia because it is the HPHPA and/or 4-cresol that are neurotoxic.

People sometimes assume that a test using DNA is more accurate than other types of testing. However, DNA testing is fraught with complexities. The nucleic acids of Clostridia are extremely diverse. The content of the nucleic acid bases guanosine and cytosine (G+C) is used to classify bacteria species. The G+C content of DNA of Clostridia species ranges from 21-54 % (9). The majority of intestinal species have G+C contents in the lower half of this range. Ribosomal RNA cataloging confirms that Clostridia occupy six independent sublines with multiple branches including non-Clostridia species. The failure to offer documentation on which species are being detected and how validation was performed should lead to caution by the user of such testing, especially when such tests may be labeled "experimental". Similar complexities exist with traditional culture methods for Clostridia since results are commonly reported from 0 to 4+. Since many Clostridia are not pathogenic, what does a high Clostridia level of 4+ indicate since beneficial, neutral, and harmful species are lumped together in one category? In reality, the results of stool tests for total Clostridia are virtually meaningless and may lead to inappropriate patient treatment.

It is estimated that there are about 10 billion cells of Clostridia per gram of stool. Clostridium ramosum is the most common (53% of all subjects tested) with a mean count of about one billion per gram of stool (9). The prevalence of some Clostridia species is highly dependent on diet. Stool samples of vegetarians did not contain Clostridium perfringens whereas meat and fish eaters had high amounts (10).

Since HPHPA is associated with multiple species of Clostridia but not Clostridium difficile, there is really no available confirmation test for determining the specific species of Clostridium producing HPHPA. As mentioned above, stool testing for total Clostridia is useless since it cannot currently differentiate between harmful or beneficial species. Since HPHPA, in my experience, disappears after treatment with vancomycin or metronidazole, I always recommend treatment based on the HPHPA value with a follow-up test 30 days after completion of treatment.

Confirmation testing of Clostridium difficile could be performed when 4-cresol is elevated. However, the prevalent testing for Clostridium difficile toxins A and B are focused on strains that cause gastrointestinal damage. Strains that produce 4-cresol but not toxins A or B may still cause significant psychiatric disease, so performing these toxin tests may muddy the interpretation of the clinical situation if these tests are negative. I think that it is easier to treat based on the 4-cresol results and then do follow-up testing of the 4-cresol on the organic acid test 30 days after completion of treatment.

Clinical References

• Beatty, H. Botulism. In: Harrison's Principles of Internal Medicine, 10th edition, ed. R. Petersdorf, et al. McGraw Hill. New York. 1983. Pages 1009-1013.

• Meyer, K.F. and Lang, O.W. A highly heat-resistant sporulating anaerobic bacterium: Clostridium caloritolerans, N. SP. The Journal of Infectious Diseases Vol. 39, No. 4 (Oct., 1926), pp. 321-327

• Chalmers, R.A., Valman. H.B., and Liberman, M.M., Measurement of 4-hydroxyphenylacetic aciduria as a screening test for small-bowel disease. Clin Chem 25:1791, 1979

• Carrico, R.M. Association for Professionals in Infection Control and Epidemiology (APIC) Implementation Guide to Preventing Clostridium difficile Infections http://apic.org/Resource_/EliminationGuideForm/59397fc6-3f90-43d1-9325-e8be75d86888/File/2013CDiffFinal.pdf (accessed Oct 30,2014)

• Sivsammye, G. and Sims, H.V. Presumptive identification of Clostridium difficile by detection of p-cresol (4-cresol) in prepared peptone yeast glucose broth supplemented with p-hydroxyphenylacetic acid. J Clin Microbiol. Aug 1990; 28(8): 1851–1853.

• Phua, T.J., Rogers, T.R., and Pallett, A.P. Prospective study of Clostridium difficile colonization and paracresol detection in the stools of babies on a special care unit. J. Hyg., Camb. (1984). 93. 17-25 17

• Yokoyama, M. T., Tabori, C., Miller, E. R. and Hogberg, M. G. (1982). The effects of antibiotics in the weanling pig diet on growth and the excretion of volatile phenolic and aromatic bacterial metabolites. The American Journal of Clinical Nutrition 35, 1417-1424.

• Persico, A.M. and Napolioni, V. Urinary p-cresol (4-cresol) in autism spectrum disorder. Neurotoxicology and Teratology 36 (2012) 82–90

• Wells, J.M. and Allison, C. Molecular genetics of intestinal anaerobes. In: Human Colonic Bacteria. Role in Nutrition, Physiology, and Pathology. Gibson and MacFarlane, ed. CRC Press. Ann Arbor. 1995. Page28

• 10. Conway, P. Microbial ecology of the human large intestine. In: Human Colonic Bacteria. Role in Nutrition, Physiology, and Pathology. Gibson and MacFarlane, ed. CRC Press. Ann Arbor. 1995. Pages 1-24

A Primer on Natural Antifungal Agents: Evidence and Rationale for Their Use

Jessica Bonovich RN, BSN

Guidelines for the treatment of yeast have been documented in the literature for nearly every major organ system (Pappas). Yet, a standard of care for gastrointestinal yeast treatment is surprisingly absent despite the large body of work demonstrating that pathogenic yeast causes harm to various aspects of the gastrointestinal tract (Zwolinska, Brzozowski). Clinicians suspicious of GI yeast overgrowth typically perform a fecal analysis with culture and sensitivity. While this method is ideal for the effective treatment of yeast, it is poorly understood why patients with yeast overgrowth often test negative upon laboratory examination of stool (Maaroufi, Shaw). Up to 50% of stool analysis negative for yeast species returned positive on PCR (Maaroufi). Metabolites of yeast detected in The Great Plains Laboratory Organic Acids Test are a very reliable method of detecting yeast (Shaw). However, this test cannot determine the exact organism and therefore its susceptibility to antifungals (Shaw). It also cannot determine the exact location of the yeast overgrowth but clinical experience has shown that the majority of cases are in fact GI related. The documentation set forth is based broadly on in vivo and in vitro studies on the antifungal properties of the natural agents, documentation of yeast infections involving organ systems other than the GI tract, and yeast overgrowth in the GI of the irritable bowel patient population.

Probiotic Support

Evidence:

Candida: Promising data by several small studies has demonstrated the use of probiotics as effective against numerous pathological conditions caused by Candida. In these studies, Lactobacillus GG, L. acidophilus, and Saccharomyces boulardi were the predominant probiotics shown to be effective with L. GG demonstrating the ability to induce antibody formation against Candida in immune deficient mice. Probiotics have been shown to accelerate the healing of various pathological conditions in the gastro-intestinal tract when Candida is present (Zwolinska 2006 & 2009, Hatakka). Probiotics have also been shown to accelerate immune response to Candida in several murine simulations (Wagner, Zwolinska 2006 & 2009).

Aspergillus: Data on the effectiveness of probiotics against Aspergillus infection is not available. Aspergillus infections are thought to be rare in comparison to other yeast species such as Candida. However, a recent study indicated a high percentage of Aspergillus in stool samples of patients with Crohn's disease. (Li) Aspergillus infections are usually associated with pulmonary infection and or post-surgical complications that are often very acute. The severity of the Aspergillus complications and small numbers of infection are presumably responsible for the lack of research in this regard. The relative safety of Lactobacilli, bifidobacteria, and lactococci has been demonstrated extensively in the literature. Incorporation of these probiotics into a protocol for Aspergillus treatment may be considered appropriate in many cases.

Studies on the use of probiotics for gastrointestinal healing have been aimed at a wide range of populations. To date, the most promising studies have been in the treatment and prevention of acute infectious diarrhea, viral gastroenteritis, antibiotic associated diarrhea, ulcerative colitis, and necrotizing enterocolitis in preterm infants (Manzoni, Zwolinska, Szajewska). In all of these conditions, inflammation is of a primary concern.

Risks: Reports of bacteremia and even a few isolated cases of sepsis have been documented in the literature from the Lactobacillus genera including L. rhamnosis, L. plantarum, L. casei, L. paracasei, L. salivarius, L. acidophilus (Snydman, Borriello). Cases of sepsis have also been documented for the usually beneficial yeast Saccharomyces boulardi. In some cases, the cultures were linked to a probiotic supplement, in others, the bacteria were found to be intrinsic to the patient's own microflora (Snydman, Borriello). In all of the cases, the patients were severely immunocompromised and often had feeding tubes, short gut syndrome, and/or a central line (Snydman, Borriello, Munoz, Herbrecht). The cases of sepsis have most commonly been associated with S. boulardi (Munoz, Herbrecht). However, fungemia from S. boulardi infection is rare in comparison to the population believed to be taking the supplement (Herbrecht, Munoz). In one study, increase in bacteremia from Lactobacillus did not increase over a decade, despite the 6 fold increase in probiotic use (Borriello). These data indicate that individuals taking probiotics are not at any greater risk than the general population for bacteremia associated with Lactobacillus. Regardless, the practitioner should exercise caution in severely immunocompromised patient populations to reduce any risk to the patient.

Rationale:

Promote the immune response against intestinal yeast overgrowth. To promote healing and reduce inflammation in the intestinal mucosa during yeast overgrowth.

Dosing:

The strain most commonly championed in the literature is that of Lactobacillus GG in doses of 10 billion colony forming units (CFU's) taken early in treatment. Saccharomyces thermophilus and S. boulardi were found to be effective in some studies and less effective in others. A daily intake of 10^6 to 10^9 CFUs is reportedly the minimum effective dose for therapeutic purposes.

Allicin

Evidence:

Allicin is the active ingredient found in garlic. The most commonly understood mode of action for allicin is linked to its ability to cross cell membranes and combine with sulfur-containing molecular groups in amino acids and proteins, thus interfering with cell metabolism (Davis, Singla). The antimicrobial properties of allicin have been demonstrated in numerous in vitro and in vivo murine models (Davis, Guo, Shadkchan). The antifungal properties of allicin have been shown to potentiate the effectiveness of fluconazole, the synergistic combination being the most effective at killing Candida species in kidney cells (Guo). Human studies have been targeted largely toward cardiovascular and antihypertensive effects and little has been done to demonstrate the antimicrobial properties (Fugh-Berman, NACAM). However, a study in China reports successful use of intravenous allicin against invasive fungal infections (Davis).

The strength of the supplement is affected by the preparation of garlic. Studies have shown that water, oil, and high temperatures can degrade allicin content (Singla). Interestingly powdered garlic is found to be the highest in allicin (Singla). Interestingly, powdered preparations of garlic for cooking were found to have a greater allicin content than nine supplement tablets studied (PDR). There are also pure allicin extracts available on the market for use.

Risks: Studies have demonstrated that allicin can inhibit platelet aggregation in blood and several cases of bleeding complications have been documented. All of which were following an invasive procedure (Fugh-Berman). Allicin may also increase production of insulin by pancreatic cells causing the potential for hypoglycemia in some patient populations. Allicin may also inhibit cholesterol synthesis in the liver causing exacerbation of developmental delay in children with low cholesterol levels. Physicians should use caution in patients with bleeding conditions, on blood thinners, with hypoglycemia, or diabetics who are insulin dependent. Cholesterol testing is advised for children with developmental disorders prior to supplementation with allicin.

Rationale:

Mild antifungal therapy when prescriptive agents are unavailable or contraindicated and where dosing by weight is required (such as for children). Promote the synergistic modulation of antifungal therapy with fluconazole.

Dosing:

Insufficient evidence exists in US literature for dosing recommendations, especially for children. However, there are several governing bodies outside of the US that regulate supplementation and provide a guideline for dosing. According to the National Center for Complementary and Alternative Medicine in the US, allicin is considered safe for most adults. Use of allicin for antifungal treatment may be appropriate in doses as high as one milligram per kilogram of body weight. Human studies have demonstrated that doses of allicin effectively potentiated the effects of antifungal treatment in doses of 7.8 - 27 mg per dose. The European Scientific Cooperative on Phytotherapy (ESCOP) recommends 3 to 5 milligrams allicin daily (1 clove or 0.5 to 1.0 gram dried powder) for the prevention of atherosclerosis. The World Health Organization (WHO) recommends 2 to 5 grams fresh garlic, 0.4 to 1.2 grams of dried powder, 2 to 5 milligrams oil, 300 to 1,000 milligrams of extract, or other formulations that are equal to 2 to 5 milligrams of allicin daily. The European Scientific Cooperative on Phytotherapy (ESCOP) recommends 2 to 4 grams of dried bulb or 2 to 4 milliliters of tincture (1:5 dilution in 45% ethanol), by mouth three times a day for upper respiratory tract infections.

MCT Oil/ Caprylic Acid/Monolauren/Coconut Oil

Evidence:

There are numerous in vitro and in vivo animal studies that demonstrate the effectiveness of coconut oil and/or its medium chain fatty acid constituents (Caprylic Acid, Capric Acid, and Lauric Acid) against Candida and other pathogens (Bergsson, Batovska, Huang, Dayrit). Human trials are much more limited. Therefore the evidence for treating yeast with this substance is based on the clinical observation of physicians who commonly treat yeast conditions. Physicians who routinely treat patients for Candida report very good success with using MCT oil/Caprylic acid. In his book, The Yeast Connection, Dr. Crook sites numerous examples of physicians who have reported this supplement as clinically useful (Crook).

Immunomodulating Properties: Like Omega-3 fatty acids, MCT's produce fewer inflammatory eicosanoids of the two- and four-series (Wan). Several in vivo studies have demonstrated anti-inflammatory properties of MCT oil and antipyretic and analgesic properties have also been documented (Canela, Intahphuak). In vivo MCTs may reduce intestinal injury and protect from hepatotoxicity which is a concern in patients taking fluconazole and itraconazole antifungal therapy (Berit, Kono). Human studies are few but promising as many of the studies are on severely immunocompromised patients who require total parernteral nutrition (TPN) and the HIV/AIDS patient population (Wanke, Dayrit, Craig, Wolfram, Chen). This patient population has responded well to the addition of MCT's. The degree to which these results apply to the general population is unclear. However, the safety of this supplement can be inferred given its effective use in severely immune compromised patient populations.

Risks: Acute toxicity tests conducted in several species of animal demonstrate that MCTs are essentially non-toxic. Ninety-day toxicity tests did not result in notable toxicity, whether the product was administered in the diet up to 9375mg/kg body weight/day or by intramuscular injection (up to 0. 5ml/kg/day, rabbits). Levels of up to 1g/kg/day have been confirmed safe in several clinical human trials (Traul). The use of MCT is only contraindicated in patients with impaired states of fat metabolism such as ketosis, acidosis, and cirrhosis (Bach).

Rationale:

Mild antifungal therapy when prescriptive agents are unavailable or contraindicated and where dosing by weight is required (such as for children).

Dosing:

Caprylic acid: PDR for nutritional supplements indicate dose as 300-1200 mg daily
Monolauren: 240 – 720 mg three times daily (adults)
Virgin coconut Oil: 2 ml/kg/day of virgin coconut oil in children
MCT: levels of up to 1g/kg/day have been confirmed safe in several clinical human trials.

Clinical References:

• Bach, AC., Babayan, VK. (1982). Medium Chain Triglycerides: un update. American Journal of Clinical Nutrition, 36(5); 950-962

• Batovska, D., et al. (2009). Antibacterial study of the medium chain fatty acids and their 1-monoglycerides: individual and synergistic relationships. Polish journal of Microbiology, 58(1); 43-7.

• Bergsson, G., et al. (2001). In vitro killing of Candida albicans by fatty acids and monoglycerides. Antimicrobial Agents and Chemotherapy, 45(11); 3209-12.

• Berit, M., Pfeuffer, M., Schrezenmeir, J., (2006). Medium Chain triglicerides. International Dairy Journal, 16(11) 1378-1382.

• Borriello, S., et al. (2003). Saftey of Probiotics that Contain Lactobacilli or Bifidobacteria. Clinical Infectious Disease, 36(6); 775-780. Doi 10.1086/368080

• Brzozowski, T., et al (2005). Influence of gastric colonization with Candida albicans on ulcer healing in rats: Effect of ranitidine, asprin and probiotic therapy. American Journal of Gastroenterology, 40(3); 286-296.

• Canani, R., et al. (2007). Probiotics for treatment of acute diarrhea in children: randomized clinical trial of five different preparations. BMJ, 335-340. Doi 10.1136/bmj.39272.581736.55

• Canela, GO., (2007). Anti-inflammatory activity of virgin coconut oil. The Philippine Journal of Internal Medicine, 45(2) 85-88.

• Craig, GB., et al. (1997). Decreased fat and nitrogen losses in patients with AIDS receiving medium chain triglyceride-enriched formula vs those receiving long-chain-triglyceride containing formula. Journal of the American Dietetic Association, 97(6); 605-11.

• Chen, FM., et al. (2005). Efficacy of medium-chain triglycerides compared with long-chain triglycerides in total parenteral nutrition in patients with digestive tract cancer undergoing surgery. The Kaohsiung Journal of Medical Sciences, 21(11); 487-94.

• Crook, W. (2000). The Yeast Connection Handbook. Jackson, TN: Woman's Health Connection.

• Davis, S. (2005). An overview of the antifungal properties of allicin and its breakdown products-the possibility of a safe and effective antifungal prophylactic. Mycoses, 48(2); 95-100. DOI: 10.1111/j.1439-0507.2004.01076.

• Dayrit, C. (2000). Coconut oil in Health and Disease: Its and Monolauren's potential as cure for HIV/AIDS. Read at the XXXVII Cocotech Metting Chennai, India July 25, 2000. http://coconutresearchcenter.org/article10526.pdf

• Fugh-Berman, A., (2000). Herbs and Dietary Supplements in the Prevention and Treatment of Cardiovascular Disease. Preventive Cardiology, 3(1); 24-32. Doi:10.1111/j.1520-037X.2000.80355.

• Guo, N., Wu, X.,, et al. (2010). In vitro and in vivo interactions between fluconazole and allicin against clinical isolates of fluconazole-resistant Candida albicans determined by alternative methods. FEMS Immunology and Medical Microbiology, 58(2); 193-201. Doi: 10.1111/j.1574-695X.2009.00620.

• Hatakka, K., et al. (2007). Probiotics Reduced the Prevalence of Oral Candida in the Elderly – a Randomized Controlled Trial. Journal of Dental Research, 86 (2); 125-130. doi: 10.1177/154405910708600204

• Herbrecht, R., Nivoix, Y., (2005). Saccharomyces cervisiae Fungemia: an adverse effect of Saccharomyces boulardi probiotic Administration. Clinical Infectious Disease, 40(11); 1635-1637. Doi 10.1086/429926

• Huang, CB., Alimova, Y., Myers, TM., Ebersole, JL. (2011). Short and medium chain fatty acids exhibit antimicrobial activity for oral microorganisms. Archives of Oral Biology, 56(7); 650-4.

• Intahphuak, S. et al. (2010). Anti-inflammatory, analgesic, and antipyretic activities of virgin coconut oil. Pharmaceutical Biology, 48(2); 151-7.

• Khodavandi, A., Alizadeh, F., et al. (2011). Comparison between efficacy of allicin and fluconazole against Candida albicans in vitro and in a systemic candidiasis mouse model. FEMS Microbiology Letters, 315. Kono, H., et al. (2003). Protective effects of medium-chain triglycerides on the liver and gut in rats administered endotoxin. Annals of Surgery, 237(2); 246-55.

• Li, Qiurong., et al. (2014). Dysbiosis of Gut Fungal Microbiota is Associated with Mucosal Inflammation in chrohn's Disease. Journal of Clinical Gastrointerology, 48:513-523.

• Maaroufi, Y., Heymans, C., De Rune, J., Duchateau, H. (2003). Rapid Detection of Candida albicans in Clinical Blood Samples by Using a TaqMan-Based PCR Assay. Journal of Clinical Microbiology, 41; 3293-3298.

• Manzoni, P., et al. (2006). Oral Supplementation with Lactobacillus casei Subspecies rhamnosus Prevents Enteric Colonization by Candida Species in Preterm Neonates: a Randomized Study. Clinical Infectious Diseases, 42(12); 1735-1742. doi: 10.1086/504324

• Munoz, P., et al. (2005). Saccharomyces cerevisiae fungemia: and emerging infectious disease. Clinical Infectious Disease, 40(11); 1625-34.

• National Center for Complimentary and Alternative Medicine (NCCAM). Herbs at a Glance: Garlic. Retrieved on 3/3/2014 from http://nccam.nih.gov/health/garlic/ataglance.htm

• Snydman, D. (2008). The Safety of Probiotics. Clinical Infectious Diseases, 46(2); S104-S111. Doi 10.1086/523331

• Pappas, P., et al. (2004). Guidelines for Treatment of Candidiasis. Clinical Infectious Diseases. 38; 161-189

• Shadkchan, Y., Shemesh, E., et al. (2004). Efficacy of allicin, the reactive molecule of garlic, in inhibiting Aspergillus spp. In vitro, and in a murine model of disseminated aspergillosis. Journal of Antimicrobial Chemotherapy, 53; 832-836. doi: 10.1093/jac/dkh174.

• Shaw ,W., (2008) Biological Treatments for Autism and PDD. Publisher: Author.

• Singla, V., Bhaskar, R, (2011). Garlic: a review. International Journal of Drug Formulation, 2. Retrieved from http://www.ordonearresearchlibrary.org/Data/pdfs/IJDFR80.pdf

• Szajewska, H., Skorka, A., Dylag, M. (2006) Meta-analysis: Saccharomyces boulardii for treating acure diarrhea in children. Alimentary Pharmacology and Theraputics, 25(3); 257-264. Doi 10.1111/j.1365-2036.2006.03202.x

• Szajewska, et al., (2006). Probiotics in Gastrointestinal Diseases in Children: Hard and Not-So-Hard Evidence of Efficacy. Journal of Pediatric Gastroenterology and Nutrition, 42(5); 454-475. Doi 10.1097/01.mpg.0000221913.88511.72

• Wagner, D., et al. (2000). Effects of probiotic bacteria on humoral immunity to Candida albicans in immunodeficient bg/bg-nu/nu and bg/bg-nu/+ mice. Journal of Microbiology/Immunology, 17; 55-59. Pdf http://www.reviberoammicol.com/2000-17/055059.pdf

• Traul, KA., Driedger A., Ingle DL., Nakhasi, D. (2000). Review of the toxicologic properties of medium-chain triglycerides. Food and Chemical Toxicology, 38(1), 79-98.

• Wan, JM., TEO, TC., Babayan, VK., Blackburn GL. (1988). Invited Comment: lipids and the development of immune dysfunction and infection. Journal of Parenteral and Enteral Nutrition, 12(6 supplment); 43s-52s.

• Wanke, CA., et al. (1996). A medium chain triglyceride-based diet in patients with HIV and chronic diarrhea reduces diarrhea and malabsorbtion: a prospective, controlled trial. Nutrition, 12(11-12); 766-71.

• Wolfram, G., (1986). Medium Chain Triglicerides for Total Parenteral Nutrition. World Journal of Surgery, 10; 33-37.

• Zwolinska, M., et al. (2009). Effect of Candida colonization on Human Ulcerative Colitis and the Healing of Inflammatory Changes of the Colon in the Experimental Model of Colitis Ulcerosa. Journal of Physiology and Pharmcology, 60(1); 107-108. Pdf http://jpp.krakow.pl/journal/archive/03_09/pdf/107_03_09_article.pdf

• Zwolinska-Wcislo, M., et al. (2006). Are probiotics effective in the treatment of fungal colonization of the gastrointestinal tract? Eperimental and clinical studies. Journal of Physiology and Pharmcology, 57(9); 35-49. Pdf http://www.jpp.krakow.pl/journal/archive/11_06_s9/pdf/35_11_06_s9_article.pdf

Usefulness of HPHPA marker in a wide range of neurological, gastrointestinal, and psychiatric disorders

William Shaw, Ph.D.

The dysbiosis marker 3-(3-hydroxyphenyl)-3-hydroxypropionic acid (HPHPA), the predominant dihydroxyphenylpropionic acid isomer in urine, is also measured in the Organic Acids Test offered by The Great Plains Laboratory. This marker was proven by Dr. William Shaw to be due to a combination of human metabolism and the metabolism by a group of Clostridia species, including but not limited to C. difficile. 

HPHPA has been one of the most useful clinical markers in recent medical history. Treatment with metronidazole, vancomycin, or high doses of probiotics of individuals with high urinary values has led to significant clinical improvements or remissions of psychosis.

The biochemical role of Clostridia in altering brain neurotransmitters is due to the fact that Clostridia metabolites inactivate dopamine beta-hydroxylase, leading to an excess production of brain dopamine and reduced levels of the neurotransmitter norepinephrine. Excess dopamine is associated with abnormal or psychotic behavior. This imbalance can be demonstrated in the Organic Acids Urine Test by observing the ratio of the major dopamine metabolite, homovanillic acid (HVA), to that of the major norepinephrine metabolite, vanillylmandelic acid (VMA) when the Clostridia marker HPHPA is elevated. After treatment with metronidazole or vancomycin, HPHPA values return to normal along with normal ratios of HVA/VMA and normal behavior. 

The highest value of HPHPA was measured in the urine of a young woman with first onset of schizophrenia. Treatment of Clostridia bacteria resulted in loss of auditory hallucinations. In autism, children with gastrointestinal Clostridia commonly exhibit aggressive behavior, agitation, obsessive compulsive behavior, and irritability. They may have very foul stools with diarrhea with mucus in the stools although some individuals may be constipated. Stool testing for Clostridia is usually of limited usefulness since most Clostridia species are considered probiotics or beneficial. There are about 100 species of Clostridia that are commonly found in the gastrointestinal tract. Only seven of these species are producers of HPHPA including C. sporogenes, C.botulinum, C. caloritolerans, C. angenoti, C. ghoni, C.bifermentans, C. difficile, and C. sordellii while C. tetani,C. sticklandii, C. lituseburense, C. subterminale, C.putifaciens, C. propionicum, C. malenomenatum, C.limosum, C. lentoputrescens, C. tetanomorphum, C.coclearium, C. histolyticum, C. aminovalericum, and C.sporospheroides do not produce compounds that are converted to HPHPA.

The same article by Dr. Shaw indicates that 3,4-dihydroxyphenylpropionic acid (DHPPA) is a marker for beneficial bacteria in the gastrointestinal tract such as Lactobacilli, Bifidobacteria, and E. coli. The exception is one species of Clostridia orbiscindens that can convert the flavanoids luteolin and eriodictyol, that occur only in a relatively small food group that includes parsley, thyme, celery, and sweet red pepper to 3,4-dihydroxyphenylpropionic acid. The quantity of C. orbiscindens in the gastrointestinal tract is negligible (approximately 0.1% of the total bacteria) compared to the predominant flora of Lactobacilli, Bifidobacteria, and E. coli (7). DHPPA is an antioxidant that lowers cholesterol, reduces proinflammatory cytokines, and protects against pathogenic bacteria. 2,3-Dihydroxyphenypropionic acid, a different isomer has been claimed to be a metabolite of Pseudomonas species but the literature indicates that this compound is formed by the in vitro action of these species on quinoline, a component of coal tar, a substance missing from the diet of virtually all humans. 

References:

1. Shaw W. Increased urinary excretion of a 3-(3-hydroxyphenyl)-3-hydroxypropionic acid (HPHPA), an abnormal phenylalanine metabolite of Clostridia spp. in the gastrointestinal tract, in urine samples from patients with autism and schizophrenia. Nutr Neurosci. 2010 Jun;13(3):135-43.