Yesterday morning a story made its way through the British press about a probiotic, newly available commercially, which, in a 2016 clinical trial, successfully alleviated stress, increased feelings of happiness and boosted memory. The probiotic, Zenflore, is being called the first commercially available “psychobiotic”: that is, a probiotic specifically designed with “potential mental health benefit.”[i]
(To call it the first psychobiotic is reaching for it, in my opinion, by the way. There are many probiotics on the market that have been shown to have a beneficial psychological effect. For example, the Lactobacillus casei strain Shirota, found in the yogurt/kefir brand, Yakult, prevented stress-related cortisol increases and raised serotonin levels in stressed medical students, according to a 2016 study, and more than that: “…the rate of subjects experiencing common abdominal and cold symptoms and total number of days experiencing these physical symptoms per subject were significantly lower in the LcS group than in the placebo group during the pre-examination period.” [ii])
Still, the clinical study – although not a cross-over one – makes this news worth reporting.
The product contains a particular strain of Bifidobacterium – Bifidobacterium longum 1714 – and it apparently increases “…activity in the areas of the brain associated with emotions, learning and memory.”[iii]
At baseline, 22 healthy volunteers completed a cognitive assessment, and also had EEGs. They were then subjected to a “cold pressor test” (which apparently causes some degree of pain) to measure baseline readings, and then after 4 weeks of taking either a placebo or the probiotic, the test was re-administered. The rise in cortisol (from the stress) and anxiety levels were measured, and in the probiotic group, these were significantly lower. More than that, daily stress was reported to be lowered. The researchers assessed cognitive performance on “…tests assessing memory, sustained attention, social cognition and emotional processing.” They also measured brain activity in various regions using EEG.
The results showed that this strain of probiotic “…can ameliorate both the physiological and psychological response to an acute stressor, as well as longer-term daily self-reported psychological stress, in health human adults.” They also found a “subtle” improvement in visual-spatial memory, and the EEGs of the experimental group supported improved memory activity in the brain.
What amazes me is that the dose of probiotic is remarkably low – only 1 billion organisms. I double checked and yes, 1 pill a day with that amount is all that is recommended and all that was used in the clinical study.
Currently, the product is only available through pharmacies located in limited European countries which must special order it. In case any of you outside those countries are interested, I took a look around to see if there are any options. One of my favorite bloggers, Dr. Paul Whiteley, who writes “Questioning Answers” (which covers scientific research on autism), states in a 2016 post that covered this study, that VSL 3 also contains this particular strain of B. longum.[iv] I cannot figure out where he got that information, but I trust him. (He is a very highly regarded blogger.) And over the years, as I have mentioned before, I have had tremendous success with VSL 3 in both the IBS/IBD and autism populations. I personally take it daily. Other companies, like Life Extension, make a Bifido Longum product but the strain is different and thus, I can’t be sure it would have the same effect.
If any of you in Europe do try Zenflore, I (and I am sure my readers) would love to hear from you about your results!
[i] Allen, AP, Hutch, W, Borre, YE, Kennedy, PJ, Temko, A, Boylan, G, Murphy, E, Cryan, JF, Dinan, TG, Clarke, G. Bifidobacterium longum 1714 as a translational psychobiotic: modulation of stress, electrophysiology and neurocognition in healthy volunteers. Translational Psychiatry. 2016: 6; e939.
[ii] Kato-Kataoka, A, et. al. Fermented milk containing Lacobacillus casei strain Shirota prevents the onset of physical symptoms in medical students udner academic examination stress. Beneficial Microbes. 2016:7(2);153-6. doi: 10.3920/BM2015.0100
Since eating a healthy diet is the most important factor in maintaining a healthy and happy inner biome, I thought I’d catch you up on some of the latest papers out there.
I spotted another “most popular medical stories of 2018” article on Medical Express[i], and took a skim through. One of the items stated, “…a combined team of researchers from the University of Aberdeen and the Chinese Academy of Sciences announced that fat consumption is the only cause of weight gain. In their study with mice, they found that sugar and carbohydrates did not lead to weight gain no matter how much was consumed.” Whoa! The low-carb/Atkins/Paleo/ketogenic diet communities must have freaked when reading that statement!
The controversy over the optimal diet for health, weight loss, etc. rages on year after year. I have read books and scientific articles about all these different diets and at this point, truly have absolutely no idea which is correct. There is scientific evidence to be found on all sides. But what I do know is that we Americans, and people from other industrialized countries around the world, are doing nothing but getting fatter. We are doing something wrong…and I have to believe in the truth of the mounting evidence that biome depletion is a factor.
In the meantime, pretty much weekly, another article is published showing the benefits of eating a Mediterranean type diet, high in fruit, vegetables, nuts, seeds, olive oil and low in dairy, sweets and lean meats. A month ago, a study[ii] was published by a team of researchers from Brigham and Women’s Hospital, Harvard Medical School, and the Harvard T.H. Chan School of Public Health which “…found a 25 percent reduction in the risk of cardiovascular disease among study participants who consumed a diet rich in plants and olive oil and low in meats and sweets.” The research was based on data from 25,000 female health professionals who provided both information about their diets and blood samples, to follow biomarkers, for 12 years.
10 days or so after I read about that, another paper[iii] came out showing, yet again, that eating junk food (foods that are high in cholesterol, saturated fats and carbohydrates) increases the risk of depression by about 40%. In this case, the scientists analyzed data from 11 existing studies covering in total, about 100,000 people. “An anti-inflammatory diet—containing more fibre, vitamins (especially A, C, D) and unsaturated fats—has the opposite effect, and could be implemented as a treatment for depression. Therefore, a Mediterranean diet of olive oil, tomatoes, green vegetables and fatty fish could help lower depressive.”
And then last week, I read about a 3rd study[iv] that demonstrated that eating more fruit and vegetables reduces the risk of memory loss. Again, data provided by 28,000 health professionals, over the course of 20 years, was used. “Researchers found that the more vegetables, fruit, and fruit juice the men consumed, the better their memory skills. Those that ate a lot of leafy green vegetables, vegetables high in carotenoids, and berry fruits lowered their risk of memory loss. Also, the men who drank more orange juice in their seventies scored better on the memory skills questionnaire.” In fact, the orange juice consumption so stood out that the researchers are recommending that this link be pursued in further research.
Of course all these studies have their flaws, but when taken as a whole – these, plus the hundreds of others in the literature – demonstrate pretty clearly that a healthy diet has tremendous effect on long-term health. As this last article concludes, “It is empowering to know that by simply eating more fruits and vegetables, and drinking orange juice can lower the risk for memory loss. This study suggests that individuals can improve brain health by making a few simple diet changes that can pay off in the long run.”
Of course, a diet rich in fiber – as I have written about over and over – is incredibly beneficial to your gut bacteria. So, for the sake of your and your gut buddies, commit to a healthy diet. It’s the best New Year’s resolution you can make for yourself.
A few days ago, a fun article[i] appeared on Gut Microbiota for Health which listed their most popular articles in 2018. Contrary me: I looked through all 10 and found that the last one on the list was way the most interesting.
In your small intestine, there is lymph tissue (GALT = gut-associated lymphoid tissue), where your cells interact with both gut microbes and immune cells. This is where the decision is made whether or not you will tolerate something in the gut (or attack it as an invader). In the GALT, gut bacteria help make an immune cell that “…enables its host to function normally by tolerating harmless substances from the outside.”
Researchers from Washington University in St. Louis’ School of Medicine wanted to find out specifically which commensal bacteria was responsible for the production of this immune cell, known as DP IEL (double-positive intraepithelial lymphocyte), which help the body to distinguish self from non-self and bad from good, regulating inflammatory responses accordingly.[ii] These cells are absent in germ-free mice, so it was already established that commensal bacteria were crucial in its production…but which specific bacteria were responsible was unknown.
By giving germ-free mice different strains of Lactobacillus and Bacteroides, these scientists were able to establish which led to increased levels of DP IEL, and they discovered two crucially important things. Firstly, the essential amino acid (meaning that you must eat it to get it – your body cannot produce it on its own), L-tryptophan, is necessary to in the process of creating DP IEL. The higher the level of tryptophan fed to the mice, the higher their level of DP IELs. (Foods that are high in tryptophan include turkey, salmon, eggs, cheese, nuts, seeds.) Secondly, they discovered that the probiotic bacteria, L.reuteri, was also crucial in its production. Interestingly, L.reuteri’s effectiveness in the production process was enhanced when they added other beneficial species to the mix.[iii]
Holy cow! How astounding is that? I have written before about how essential L.reuteri seems to be in illnesses like autism and PTSD, both of which are associated with alterations in the microbiome and high levels of inflammation. And by the way, tryptophan metabolism, which is the key in the formation of messenger molecules like serotonin, has long been considered suspect in various “mental” illnesses. This tryptophan/L.reuteri combo has not as yet been tested in humans, but I am sure that’s only a matter of time. In the meantime, since we already know it has a positive effect on autism, depression and anxiety disorders, adding some to the diet is certainly not going to hurt.
[iii] Cervantes-Barragan L, Chai JN, Tianero MD, DiLuccia B, Ahern PP, Merriman J, Cortez VS, Caparon MG, Donia MS, Gilfillan S, Cella M, Gordon JI, Hsieh C-S, Colonna M. Lactobacillus reuteri induces gut intraepithelial CD4 CD8 alpha alpha T cells. Science. 2017; 357(6353): 806-810. doi: 10.1126/science.aah5825.
Remember back in September, I was ranting about two “anti-probiotic” studies and the insane conclusions people were drawing based upon them? Well, a few days ago, I came across an article[i] on this very topic, on the Gut Microbiota for Health website, that is worth taking a few minutes to write about. (This site was created by the Gut Microbiota and Health Section of the European Society for Neurogastroenterology & Motility, a scientific organization, and is devoted to posting the latest microbiome science.)
It turns out, it wasn’t just me who was beyond annoyed. In this case, the pissed off people are scientists who study the microbiome and who are fed up with the recent fake news “anti probiotics” campaign the media has been waging: “A few weeks ago, one piece of news hit the headlines and caused quite a commotion. Stating that probiotics were little more than ‘useless’ and could even have negative effects on health, this information was based on a couple of pieces of research published in the scientific journal Cell.”
A few examples:
Dr. Lorenzo Morelli, director of the Science and Food Technology Department at the Università Cattolica del Sacro Cuore (Italy), states, ““We have a large number of pathogenic bacteria in the gut, and when there is an imbalance between these and our good bacteria, health problems emerge. Probiotics can help avoid this imbalance and, consequently, limit the consumption of antibiotics.”
Dr. Patrice Cani, a researcher at the Belgian Fund for Scientific Research (who studies a subject near and dear to us Biome Buzzers – the role of the Akkermansia muciniphila bacteria in obesity, diabetes and heart disease), points out that not all probiotics are the same, or will have the same effect in people. “Even if the bacteria are only passing through the intestine, they can produce active compounds or induce the immune system to produce other, specific antimicrobial factors ‘simply’ through contact with the mucus.”
Dr. Cani also points out that “…not all bacteria have a positive effect on human health, and neither are all probiotics the same and useful for everything. In other words, the properties of one cannot be attributed to others.”
Gut Microbiota For Health was much tamer than feisty me in its critique of the media hype, but the message was the same: do not write off probiotics’ well-documented health benefits based upon two tiny studies. “Over more than a decade, clinical trials involving thousands of patients have demonstrated that probiotics are beneficial to health.”
It’s not often I find stories about any of the other “-omes” outside the microbi-ome, so I was pretty excited to find a big piece of news this week related to the virome.
Microbiologists at Princeton University discovered a phage (remember, bacteriophages are viruses that kill bacteria) that actually spies on the conversations of bacteria to know when the time is ripe for attack.[i]
According to the lead scientist, Dr. Bonnie Bassler, viruses can only make 1 decision: to either remain inside the host “under the radar,” or to replicate itself and attack other nearby hosts. If the virus attacks too soon and there are not enough viable hosts in close enough proximity, the virus and its offspring will just die.[ii]
These researchers though have discovered that this particular phage (VP882) has evolved to “listen” to the chemical crosstalk between bacteria, signaling that they are in a crowd.
Dr. Blasser discovered years ago that bacteria communicate and sense each other’s presence, so that they know there are enough of them to mount a successful attack. Discovering that a bacteriophage can spy on them and understand that message, so that it too could pick an optimal time for its own attack, was a stunning discovery: this virus remains dormant until bacteria have reproduced enough to mount an attack, and then it swoops in, destroying the invading bacterial army.
What’s truly remarkable about this is that these are radically different organisms, from different biological kingdoms. The fact that this virus can understand the “language” of another species is astounding. To boot, while most viruses can only attack 1 particular kind of cell (for example, the flu virus only infects lung cells), so far VP882 has shown it can recognize, and optimize an attack on, cholera, salmonella and E.coli.
This is a huge step forward in creating therapeutic phages to treat bacterial infections: phages are totally targeted in their attack and thus, would spare the commensal bacteria of the gut.
Yet another article[i] was just published showing that dealing with inflammation issues while a baby is still in the womb can make a huge difference on the risk of atopic (allergic) illnesses.
In this case, researchers in New Zealand gave 474 pregnant (starting at 35 weeks) moms the probiotic Lactobacillus Rhamnosus (boy, I’ve been writing a lot about that lately!) and then their newborns, up until the age of 2 years. This cut the children’s rates of developing eczema in half! (They followed the children through age 11.) The probiotic also afforded some protection against the development of asthma, hay fever and allergies. This was actually sort of a replication study: in a previous study, the scientists showed that the probiotic significantly lowered eczema rates in these children through the age of 6.
The study design: the moms and then their babies were broken up into 3 groups. One received the L.rhamnosus (6 billion units), one received the probiotic, Bifidobacterium lactis (9 billion units), and the third got the placebo. The L.rhamnosus group had a 50% reduction in eczema compared to both other groups. Like the placebo, the B.lactis had no effect. (By the way, the moms also breast fed their babies. And you know, from my post last week, that the prebiotics found in breast milk are crucially important in helping these good bacteria to colonize.)
In reading an article on Medical Express which covered this research, I was shocked to learn, that 30% of children under the age of 2 in New Zealand suffer with eczema.[ii] Holy cow. I was curious if it was equally prevalent elsewhere and found that yes, it’s insanely common. 10% of the United States populace suffers from it.[iii] Rates are similar in other industrialized countries. According to AllergyUK, for example, 15-30% of children in the country suffer from eczema, dropping off to about 10% again by the time they reach adulthood.[iv]
Once again, I find myself concluding a post saying, wow – if I had known all this 25 years ago, I would have done things very differently during my 2 pregnancies.
[i] Wickens, K, et al. Effects of Lactobacillus rhamnosus HN001 in early life on the cumulative prevalence of allergic disease to 11 years, Pediatric Allergy and Immunology (2018). DOI: 10.1111/pai.12982
From the earliest days of this blog, I have talked about the too-often-ignored (and non-existent-in-most-people) macrobiome. Over the last few years, I’ve covered research papers that demonstrate over and over that helminths – intestinal worms, which are the predominant organisms of the mammalian macrobiome – are a potent stimulator of Th2 regulatory cytokines that modulate the inflammatory response.
In tidying up my desk today, I realized I have not yet told you about the one clinical trial done looking at the effects of helminths in autism spectrum disorders (ASD). This study was done several years back, but only just published.[i]
The justification for the trial lies in the fact that “Current literature supports a link between neuroinflammation, imbalanced immune responses, and ASD.” Researchers have found neuroinflammation in various parts of the autistic brain. Also, in animal models, maternal immune activation is known to result in autistic-like behaviors in offspring. Treating the pregnant animals with anti-inflammatory agents protects the babies, even when the mothers are immune activated. Thus, “…due to the inflammatory mechanisms implicated in the development and symptomatology of ASD, immunomodulatory interventions should be explored…”
An important side-note: Dr. William Parker, of Duke University Medical School, has also done research on this and shown that helminths are protective of babies. He used an animal model (rats) and the benign helminth, Hymenolepis diminuta cysticercoid: “We have demonstrated previously that rats infected with bacteria as newborns display life-long vulnerabilities to cognitive dysfunction…Here, we demonstrate that helminth colonization of pregnant dams attenuated the exaggerated brain cytokine response of their offspring to bacterial infection, and that combined with post-weaning colonization of offspring with helminths…completely prevented…cognitive dysfunction in adulthood.” By down-regulating the inflammatory response, helminths were able to protect babies’ brains from disrupted development, even when immune challenged.[ii]
After all (back to today’s paper): “The interaction of the developing immune system with microorganisms, including helminths, may be an important component of normal immune system maturation.”
As a proof of concept study, this clinical trial was small, only 10 higher functioning adults with autism. The scientists were simply looking to see how big an effect the helminths would have on repetitive behaviors, irritability and global functioning. They used TSO, Trichuris suis ova, which are whipworms native to pigs. They die in humans in about two weeks, and have “no known pathogenic potential.”
The test subjects were broken up into two groups, one receiving the TSO (2500 organisms every 2 weeks), one receiving the placebo. At the end of 12 weeks, each group went through a 4 week “wash out” period (during which they were given nothing), and then they were switched.
Considering that the 12 weeks on the helminths is not a particularly long time at all, the results were pretty darn remarkable.
“There were large effect sizes for TSO on rigidity and repetitive behaviors in ASD subjects…Multiple scales showed a significant percent change from baseline, indicating improvements in repetitive behaviors…Trending improvements were observed in irritability…”
As far as social communication, there was a small beneficial effect. I’m not sure how much improvement should have been expected in language in only 12 weeks.
The paper concludes with saying, “The potential impact of TSO vs. placebo in ASD is supported by moderate to large effect sizes….” and the authors of course suggest a future clinical trial, with a larger patient base. Good luck with that. Dr. Parker has been unsuccessfully trying to raise funds for helminth clinical trials for years. Considering that, as my regular readers know only too well, there is no treatment for autism, you would have thought this trial would have spurred huge action. Unfortunately (and tragically) not….
[i] Hollander, E, Uzunova, G, Taylor, BP, Noone, R, Racine, E, Doernberg, E, Freeman, K, Ferretti, CJ. Randomized crossover feasibility trial of helminthic Trichuris suis ova versus placebo for repetitive behaviors in adult autism spectrum disorder. World Journal of Biological Psychiatry. 2018 Nov 16:1-9. doi: 10.1080/15622975.2018.1523561.
[ii] Williamson, LL, McKenney, EA, Holzknecht, ZE, Belliveau, C, Rawls, JF, Poulton, S, Parker, W, Bilbo, SD. Got Worms? Perinatal exposure to helminths prevents persistent immune sensitization and cognitive dysfunction induced by early-life infection. Brain, Behavior and Immunity. 2016: 51:14-28. doi: 10.1016/j.bbi.2015.07.006
The article I read yesterday was a natural follow-on from my last post, about autism and its relationship to perturbations in the gut microbiome. This one was about a double-blind, placebo-controlled study using probiotics to restore normal gut microbiota in infants who were either born via c-section or given antibiotics early in life.[i]
Before I describe the study and its results, I want to list a few of the critical points the paper makes in its opening paragraphs:
This particular study was really very interesting in that these researchers tested the efficacy of this same probiotic mix in a variety of birth situations and their results are pretty incredible.
First, they looked at babies born vaginally who received no antibiotics. The levels of Bifidobacterium breve and Lactobacillus rhamnosus were significantly higher in the supplemented group (up to 10X more in the babies who were breast-fed, less though in the formula fed ones). In the control group (so not supplemented with the probiotic), breast-fed babies still had higher levels of B.breve but not even vaguely as high as those given the supplement, while the un-supplemented formula-fed babies did not have B.breve as a dominant group, “…suggesting that B.breve is a natural colonizer of breastfed infants.”
In fact, they discovered – looking further at the bacteria of these babies – that breast-fed babies had specific differences to the formula-fed ones: “…most of the supplement-induced changes observed in the breast-fed infants were not present in the formula-fed infants. In the breast-fed group, the abundance of lactobacilli was 100% increased and that of bificobacteira 29% increased in response to the supplement.” So breast milk has a huge effect on the colonization of good flora. More on this later.
The scientists went on to test whether or not, using the probiotic, they could ameliorate the alterations in gut flora in babies who were either born via c-section or given antibiotics.
The c-section babies’ microbiota was “…clearly different from the vaginally born infants.” However: “Remarkably, in the supplemented group, birth mode did not have a significant impact on microbiota composition.” Wow.
Next, they tested the effects of antibiotics on the microbiota and whether or not the probiotic could ameliorate any changes. In the probiotic-untreated control group, “…infants who had been treated with one or more antibiotic courses showed a clearly different microbiota composition compared to those that had received no antibiotics.” However, in the experimental (probiotic treated) group, “…antibiotic use did not have significant overall impact.” Double wow.
The results therefore demonstrated that “…most of the antibiotic- and caesarean-associated changes in the faecal microbiota of infants could be corrected or reduced by a probiotic supplementation to mother and infant.” Furthermore, “….breastfeeding together with the probiotic supplementation offer optimal results in terms of supporting the microbiota development in these infants.”
A couple of other items of note:
The article’s conclusion: “Early-life antibiotic treatments and caesarean birth influence a large fraction of the global population and are associated with global epidemic health problems…Therefore, treatments achieving even modest improvements at the level of individuals have the potential to induce great health benefits at the population level.”
As the parent of a son with autism, and a history of inflammatory bowel disease and immune deficiency, who was given antibiotics intravenously for 5 days starting at 36 hours old (and told to stop breast feeding), this kind of research has become tremendously personal to me. Believe me – were I pregnant now, I’d do things very differently than I did 25 years ago…
[i] Korpela, K, et. al. Probiotic supplementation restores normal microbiota composition and function in antibiotic-treated and in caesarean-born infants. Microbiome. 2018:6(182). DOI: 10.1186/s40168-018-0567-4}
Two days ago, I spotted a short commentary[i] in the journal, International Journal of Molecular Sciences, from this past August, which really caught my attention. Here is why: it was written by 3 researchers in the department of neurology at Rutgers University’s medical school who state that, “We will explore the potential for treatment of ASD [autism spectrum disorders] by targeting the microbiome with probiotics….this paper will attempt to provide significance to the aggregation of the research in this area of research.” It really is a great summary of what we now know.
Here are just some of the facts the paper presents:
They conclude this list of findings by stating, “Taken together, all these microbiome alterations may be associated with the increased gastrointestinal disturbances in individuals with ASD.”
More than that, there are other highly significant findings in autism that point to a gut origin. A few examples:
Ok – so let’s summarize. We know that people with autism have GI symptoms markedly more often than people without ASD, and that the severity of those symptoms correlate with the severity of their autism. We know that they have altered gut microbiota, including SIBO, lower levels of beneficial bacteria, lower levels of anti-inflammatory SCFAs, increased frequency of leaky gut/leaky brain. We know that “This ‘leaky gut’ theory would offer a mechanism by which GI disturbances could play a role in neurodevelopment and cognition.”
But wait…there’s more. We know that TREATING these microbiome alterations alleviates the symptoms of autism. Again, just a few of the findings the paper cites.
(Just yesterday, in fact, I read about a study[ii] out of Baylor College of Medicine wherein, researchers tested probiotics in a rodent model of autism. The researchers report that, “…administration of the bacterium Lactobacillus reuteri could lead to specific changes in the brain that reverse social deficits through a mechanism that involves the vagus nerve and the oxytocin-dopamine reward system.” (I have written about L.reuteri and autism before in a post about these researchers’ previous paper on this subject!) When the vagus nerve, which connects the brain to the gut (and many other parts of the body), is activated it releases oxytocin, a hormone that increases social behavior. Lowering the level of the L.reuteri in the guts of pups reduces their social behavior. Increasing the level restores it. If the vagus nerve that connects the gut to the brain is cut, L.reuteri does not restore social behavior, proving that it exerts its effect through the nerve. In mice who were genetically engineered to “…lack oxytocin receptors in the reward neurons or blocked the receptors with specific drugs, L.reuteri also could not restore social behaviors in the ASD mice.” Thus, the scientists were able to determine the exact mechanism by which the probiotic was exerting its effect.)
Back to the commentary, in the concluding paragraphs, the researchers say, “While research into the gut-brain connection in autism still remains in its preliminary phases, there is a convincing body of evidence that suggests a relationship between gastrointestinal distress and autism….”
A week ago, two papers[iii] were published in the eminent journal, Pediatrics, which showed that 1 in 40 children born in the United States (between the ages of 3 and 17) has autism. This is literally a national catastrophe at this point. Obviously, we need way more research into the gut-brain axis in autism. After all, as these scientists write, “…even today, it is becoming evident that the gut, specifically the disturbance of it, plays an important role in certain neurological disorders including ASD.”
[i] Fowlie, G, Cohen, N, Ming, X. The perturbance of microbiome and gut-brain axis in autism spectrum disorders. International Journal of Molecular Sciences. 2018;19(8): pii: E2251. doi: 10.3390/ijms19082251.