Journal Club: The Microbiome Autism Connection

As I’ve mentioned in other posts, scientists have to read about and understand the current scientific literature. Lots of the time this is done alone, at your desk in the office or the lab, for hours and hours so that you can really understand whatever topic it is that you’re studying. But one of my favorite ways to share scientific papers is through a weekly meeting with the whole lab called a Journal Club. Although my husband laughs about this kind of nerdy science “club” (akin to his amusement about scientific societies), it’s a great way to discuss a particular topic and dive deep into a discussion about how the researchers got their results and came to their conclusions. This is the first of many Journal Clubs where we will do an abbreviated version of what we would discuss in a typical journal club in the lab. 

Paper TItle:  Reduced Incidence of Prevotella and Other Fermenters in Intestinal Microflora of Autistic Children

Authors: Dae-Wook Kang , Jin Gyoon Park ,Zehra Esra Ilhan, Garrick Wallstrom, Joshua LaBaer, James B. Adams, Rosa Krajmalnik-Brown
Full disclosure, Dr. LaBaer is the Director of the center I previously worked in at the Biodesign Institute at ASU. Drs. Park and Wallstrom worked in offices down the hall from me and Dr. Krajmalnik-Brown was in another center at the Biodesign Institute.

Journal: PLOS One (PLOS stands for “Public Library of Science”). In case you want to read the whole article, it can be downloaded (for free) here

Background/Introduction: Before this paper was published, scientists knew that many children with autism also had gastrointestinal (GI) issues suggesting that there may be a connection between the two. There have been some studies looking at antibiotic treatment (which could change the gut microbiome) before 3 years of age and how this might be connected with autism.  There have also been studies connecting the gut microbiome and the brain. So there was evidence that the gut microbiome and autism may be related in some way.

When this paper was written, scientists also knew about the microbiome and how changes in the bacteria (all 1,000,000,000,000,000 of them) that are in the gut are found in patients with many different diseases – from C. diff infection to obesity to depression.

Goal of this paper: Look closely at the changes in the gut microbiome of children with autism to better understand how these two might be related.

Methods/what did they do?: Bacteria in fecal samples is considered representative of bacteria in the gut microbiome. Therefore, the researchers collected fecal samples from children both with (20 children) and without (20 children) autism.  The samples from patients without autism were used as a “control” to compare to the autistic samples. The researchers also asked the children (or their parents) questions to help determine the level of GI issues, the severity of their autism, and their environmental factors like their diet. The researchers isolated bacterial DNA from each of these fecal samples and then sequenced the DNA to determine what types of bacteria are in the gut.

autism microbiome diversity

A figure from the paper comparing the phylogenetic diversity (PD) of the bacteria in autistic versus non-autistic children. you can see that the red boxes (for autistic children) are lower than the blue boxes (for non-autistic children) indicating a lower microbiome diversity.

Results: Through sequence analysis and other statistical methods, the authors found that children who did not have autism have a more diverse microbiome compared to autistic children.  If there is higher diversity, it means that the gut contains more different types of bacteria, and lower diversity means a smaller variety of bacteria in the gut. They also found that in the autistic patients with a greater diversity in their microbiome, their autism was generally less severe. They also did not find any correlation between age, gender or diet with these microbiome changes.

The scientists also looked at what specific genus and species of bacteria were more represented in non-autistic versus autistic children. Specifically the bacteria from genus Veillonellacaea, Provetella, and Coprococcus  are less abundant in autistic children.

Discussion/Significance: What does this all mean?  The researchers did find a correlation between decreased gut microbiome diversity and autism. It should be clarified that just because GI problems are often found in autistic children and the severity of the GI issues correlates with the severity of autism, this does not necessary mean that GI issues cause autism or vice versa.  That still needs to be determined. Also because the diversity of bacteria in autistic children is low, it is not clear if this is a cause of autism or an effect of a child having autism.  However, this paper does provide a “stepping stone” to better understand what is happening in the gut of autistic children and may help define a target for diagnosing autism (by looking at the decreased diversity in the gut as a diagnostic test) or treatment (perhaps through fecal transplant).

What has been done since? This paper was published in 2013.  So what has changed since the paper was published?  Do we know whether or not changes in the gut microbiome cause autism or not?  Unfortunately, this is still unclear.  However, if these microbiome changes are a cause of the neurological changes in autism, then one would want to do a clinical trial to test what happens to autism symptoms when the microbiome has been altered.  This could be done in a number ways including diet modulations, prebiotics, probiotics, synbiotics, postbiotics, antibiotics, fecal transplantation, and activated charcoal.  Researchers have started this process by holding a meeting that included patients and their families to figure out how this type of trial could be designed (for more details, check out this journal article).

For more information about the microbiome/autism connection, check out Autism Speaks. To read more Journal Clubs, visit the archives here.

Five Ways for You to Participate in Science – Citizen Science

Bunsen_burner

The Bunsen burner I didn’t have. Thanks Wikipedia for the image

I had a chemistry set growing up.  It was small with tiny white bottles holding dry chemicals that sat perfectly on the four tiny shelves of an orange plastic rack.  My dad would let me use the workbench in the basement to do experiments – entirely unsupervised!! You might expect that I did really interesting chemical reactions, and this formative experience helped me to develop into the curious scientist that I am today. Completely wrong.  I remember following the instructions, mixing the chemicals, and then getting stuck because I didn’t have a Bunsen burner.  So many chemical reactions rely on heat, and the green candle stuck to the white plastic top of an aerosol hairspray can wasn’t going to cut it.

My main options for doing science as a kid revolved my failed chemistry experiments, my tiny microscope and slides, and a butterfly net that never netted a single butterfly (not for lack for trying).  However, today with computers (that’s right – no computer growing up – that’s how old I am!) there are hundreds if not thousands of ways for people to get involved in science, without having to invest in a Bunsen burner. This citizen science movement, relies on amateur or nonprofessional scientists crowd-sourcing scientific experiments. I’m talking large scale experiments run by grant-funded university-based scientists that have the possibility of really affecting how we understand the world around us. One example you may have heard about is the now defunct Search for Extraterrestrial Intelligence (SETI) which used people sitting at their computers to analyze radio waves looking for patterns that may be signed of extraterrestrial intelligence. They didn’t find anything, but it doesn’t mean that they wouldn’t have if the program had continued!

Here are five ways that you can become involved in science from where you’re sitting right now!

americangut1. American Gut: Learn about yours (or your dog’s) microbiome

For $99 and a sample of your poop, you will become a participant in the American Gut project. After providing a sample, the scientists will sequence the bacterial DNA to identify all of the bacterial genomes that are present in your gut.  This study already has over 4,000 participants and aims to better understand all of the bacteria that covers and is inside your body – called your microbiome – and to see how the microbiome differs or is similar between different people or between healthy people versus those who may be sick. The famous food writer Michael Pollan wrote about his experience participating this the American Gut project in the New York Times.  They are also looking at dogs and how microbiota are shared with family members, including our pets!

2. Foldit: solve puzzles for sciencefoldit

Puzzles can be infuriating, but at least they have a point to them when you get involved in the Foldit project.  Proteins are the building blocks of life.  Made out of long strings of amino acids, these strings are intricately folded in your cells to make specific 3D shapes that allow them to do their job (like break down glucose to make energy for the cell).  Foldit has you fold structures of selected proteins using tools provided in the game or ones that you create yourself.  These solutions help scientists to better predict how proteins may fold and work in nature.  Over 240,000 people have registered and 57,000 participants were credited in a 2010 publication in Nature for their help in understanding protein structure.  Read more about some of the results here.

3. EyeWire: Mapping the BrainEyeWire-Logo

The FAQs on the EyeWire website are fascinating because as they tell you that there are an estimated 84 billion neurons in the brain, they also insist that we can help map them and their connections. After a brief, easy training, you’re off the the races, working with other people to map the 3D images of neurons in the rat retina.  You win points, there are competitions, and a “happy hour” every Friday night. The goal is to help neuroscientists better understand how neurons connect to one another (the connectome).

4. Personal Genome Project: Understanding pgpyour DNA

The goal of the Personal Genome Project is to create a public database of health, genome and trait data that researchers can then use to better understand how your DNA affects your traits and your health. This project recruits subjects through their website and asks detailed medical and health questions.  Although they aren’t currently collecting samples for DNA sequencing because of lack of funding, they have already sequenced the genomes of over 3,500 participants. The ultimate goal is having public information on over 100,000 people for scientists to use.

mindcrowd5. MindCrowd: Studying memory to understand Alzheimer’s Disease

Alzheimer’s Disease is a disease of the brain and one of the first and most apparently symptoms is memory loss.  MindCrowd wants to start understanding Alzheimer’s disease by first understanding the differences in memory in the normal human brain.  It’s a quick 10 minute test – I took it and it was fun!  They are recruiting an ambitious 1 million people to take this test so that they have a huge set of data to understand normal memory.

This is a randomly selected list based on what I’m interested in and things that I’ve participate in, but you can find a much longer list of projects you can participate in on the Scientific American website or through Wikipedia.  Also, if you’re interested in learning more about the kind of science that people are doing in their own homes, the NY Times wrote an interesting article: Home Labs on the Rise for the Fun of Science.  If decide to try one out, share which one in the comments and what you think!

No s**t?!?! Sharing poop to cure disease – it happens, it works, and here’s why!

I was talking to my sister and four-year-old nephew the other day and my sister prompted him to tell me what he wanted to study when he grew up. He looks right at me and answers “poop”. Totally funny coming from the boy who really is obsessed with his own poop, but as a scientist, I responded that I could tell him lots about poop and asked, “what about poop are you interesting in studying?”  His response, “All of it.” Well, I agree. Poop is far more interesting than we give it credit for.  In this series of posts, I will share with you all the interesting stuff I know about poop. The first post was facts about poop and post is about using poop as a cure for diseases.  Let’s get down and dirty…

gutbalance

Changes in the percentage of different bacterial species in found in patients with various diseases compared to healthy control. From a review in Nature Reviews Microbiology

Your gut is filled with bacteria – estimates of over 100 trillion bacteria – weighing nearly 3 pounds! These bacteria are essential to help digest food and produce vitamins.  These bacteria are also protective against pathogens, like other bad infectious bacteria or viruses.  By studying the proportions of different gut bacterial species, scientists have found that the percentages of these bacterial species is different in patients with various diseases .  Although it isn’t clear if the changes in the gut mircobiome are a cause or consequence of the disease, scientists and clinicians are exploring whether changing the balance of bacteria back to normal can cure (or reduce the symptoms) of these diseases.

cdiff

C. Diff bacteria

In one particular case, it’s clear that a change in the gut microbiome is the cause of the disease and that’s Clostridium Difficile (also known as C. Diff) infection. This infectious bacteria releases toxins that cause mild but annoying symptoms like watery diarrhea 2-3 times per day with abdominal pain or tenderness, but can lead to more severe life-threatening issues like watery diarrhea over 15 times per day or creating a hole in the intestines.  C. Diff is responsible for  ~1% of all hospitalizations per year (>330,000 patients per year) and > 20,000 deaths per year and costs over $3.2 billion dollars for care. The elderly, hospitalized patients or patients taking antibiotics are most at risk for transmission, which is caused by fecal-oral transmission or through hospital workers (since the C. Diff spores aren’t killed by alcohol).

Although an annoying, deadly and expensive disease, C. Diff infection has recently gotten worse.  C. Diff infection is becoming more common and relapse after treatment is more frequent. The bacteria has become more virulent with an increased capacity to produce the symptom-inducing toxins while at the same time becoming resistant to the most common antibiotics used to treat this infection: metronidazole and vancomycin.

So what does this have to do with the microbiome? Well, the normal bacteria in your gut can protect against C. Diff infection.  C. Diff is found in 2-5% of all people who aren’t sick, because the gut microbiome can inhibit C.Diff growth or toxicity directly by making antimicrobial peptides or indirectly by creating an inhospitable environment for C. Diff to grow. Because of this, scientists thought – what if we just fixed the microbiome in C. Diff infected patients so that it’s normal again? And how would they do that? Fecal (POOP) transplant!

poopeatersAlthough you may not be aware, there is a long and storied history of using poop as a treatment – primarily though the eating of poop.  Yes, this is gross, but maybe if we use the official name for eating poop it will sound less gross?  Coprophagy is the consumption of feces, with the distinctions of heterospecific coprophagy being eating feces of other species, allocoprophagy being eating feces of other individuals, and autocoprophagy being eating your own feces. If you have a dog, you know that they love eating poop (their own, cats poop, random poop, all poop really) and so do lots of other animals.   At this point (if I haven’t lost you), you may be wondering why in the world would any animal or person eat poop?? It can help in the  development of the GI tract by helping colonize the gut with bacteria, in developing resistance to pathogens, or in obtaining nutrients. In humans the practice goes back to 300AD in China where fresh, fermented, dried or infant-derived feces, charmingly named “yellow soup”, was used to treat multiple food poisoning or severe diarrhea.  In 1696, Christian Paullini wrote a book on the medical uses of human and animal feces and in 1958 the first modern description of a fecal transplant was described to treat pseudomembranous colitis.

How does fecal transplant work today? The goal is to recolonize the patient’s gut with “normal” gut bacteria.  To get this “normal” gut bacteria, you need a donor. Donors are often family or friends of the patient who are healthy and don’t have any recent antibiotic use.  There is some testing (costing $500-$2000) required to make sure that the poop doesn’t contain particular bacteria or viruses.  Then the poop is prepared for transplant. 50-60 grams of stool is added to a liquid like saline or milk and mixed together in a blender to create a liquid slurry (often patients are requested to provide their own blender). The slurry is then filtered through a coffee filter or metal strainer to remove particulates.

nasolThe feces mix then needs to get into the patient’s gut – and this can be done in a few different ways.

  • Naso-duodenal – from the nose into the stomach using a tube. Although fast and 76% effective, it’s not palatable and often has disgusting side effects like vomiting
  • Transcolonoscopic, which invloves drizzling the fecal mixture out of a colonoscpy tube in the large and small intestines. 89% effective, this method puts the poop directly where it needs to be.
  • Enemas are also highly effective (95%) and are both cheap and safe.  It can be performed at home, but it isn’t recommended.

New methods continue to be developed for fecal transplant to decrease the “gross” factor.  The feces has been dried out and turned into pills.  This is less smelly, but requires ingestion of 24-35 capsules and is more expensive.  Scientists are also trying to culture the correct bacteria mixture in the lab so that a poop donor and poop sample preparation aren’t needed. The best part about this method is that scientists have dubbed it “rePOOPulating” the gut!  Until then, a new business of stool biobanks like OpenBiome are cropping up to meet the need of poop for fecal transplants. For $500, your doctor can request a poop sample for fecal transplant.

For everyone who is completely grossed out right now, it’s important to point out that in patients with C. Diff, this treatment has been over 90% effective in recurrent infections whereas all other treatments were less than 40% effective. Patients are completely on board for this treatment. The biggest issue has been doctors who are grossed out and getting them to use poop to treat these horrible diseases.brownforyou

So next time you look at your poop as you flush it down your toilet, remember how useful poop can be and knowing “what brown can do for you.”

No s**t?!?! Interesting facts about poop

I was talking to my sister and four-year-old nephew the other day and my sister prompted him to tell me what he wanted to study when he grew up. He looks right at me and answers “poop”. Totally funny coming from the boy who really is obsessed with his own poop, but as a scientist, I responded that I could tell him lots about poop and asked, “what about poop are you interesting in studying?”  His response, “All of it.” Well, I agree. Poop is far more interesting than we give it credit for.  In the next two posts, I will share with you all the interesting stuff I know about poop.  This post will be facts about poop and the second post will be about using poop as a cure for diseases.  Let’s get down and dirty...

fecalmatterI’m not one of those people fascinated by poop.  I have never read any of the most popular books on the topic “Everyone Poops” or “What’s Your Poo Telling You“. In fact, I won’t even admit that I poop myself (as my husband will attest I insist that it’s all butterflies and rainbows down there).  But (butt!) being in a lab makes you think about things you never expected.  A common laboratory activity is something called a journal club. Held weekly, undergrads, graduate students and post-docs take turns discussing a scientific topic or journal article.  I like talking about the newest technology and controversial topics, so when it was my turn, I decided to look into the ancient, but recently rediscovered, therapeutic uses of poop to help cure diseases. As a started my research on the topic, I realized that I knew very little about poop in general.  Being the scientist that I am, I went to learn more.  And lucky you, I’m going to share!

watering_poopFirst and foremost, what is poop made of? The majority (75%) is water! The remaining 25% is a mix.  About a third of this 25% (doing the math, that’s 7.5% of your poop) is dead bacteria (back to that later) and a third fiber and undigested food (like those corn kernels you didn’t chew before swallowing).  The final third contains living bacteria, protein, cell linings, fats, salts, and substances released from the intestines and liver. In fact, the brown color of poop comes from some of these secreted substances such as bile and also bilirubin, which comes from dead red blood cells.

seven types of poopThere are seven different types of poop that have been categorized in the Bristol Stool Form Scale (or BSF for short) developed by Dr. Ken Heaton from University of Bristol.  I was going to spend the next 5 minutes wondering exactly what sort of methodology brought him to discover this seven type system, but then I just looked at the original article. “Sixty-six volunteers had their whole-gut transit time (WGTT) measured with radiopaque marker pellets and their stools weighed, and they kept a diary of their stool form on a 7-point scale and of their defecatory frequency.” I’m glad I was not a volunteer in that study – keeping a daily diary of my stool form and have the length of time from mouth to poop tracked – ick!  However, Dr. Heaton was able to conclude that the form the stool takes depends on the time it spends in the colon, with 3 and 4 being ideal stools. Now one more thing for siblings, partners, and spouses to argue about – who’s poo is better?

But(t) let’s get serious.  Besides being an indication of intestinal health, poop is also filled with bacteria.  These bacteria are representative of the bacteria that can be found in your gut and are part of your “microbiome“. Your microbiome (all of the bacteria and other bugs in and around your body) outnumber your human cells 10 to 1, and scientists think that 300-1000 bacterial species inhabit the GI tract alone!  We’re not entirely sure exactly how many species because most of these bacteria don’t grow outside the gut (in the presence of oxygen), and when we look for gut bacteria by sequencing the DNA of poop samples, we’re not sure if the bacteria in poop represents all the bacteria that are found in the gut.

Either way, what do all those bacteria do? They help with digesting food and producing vitamins.  They regulate fat storage and do some crazy things like influence the immune system and the brain (more on that in a future post).  These bacteria are also protective against pathogens, like bad  infectious bacteria or viruses. How the gut microbiome protects against pathogens is still being studied, but we know that some gut microbiome bacteria create antimicrobials that kill bad bacteria.  In other cases, its all about the balance of the good bacteria versus the bad.  When this balance changes, it can be a cause or consequence of the disease. And one of the cures to these diseases, might just be poop itself, which is what I’ll discuss in my next post.

Want to learn more about poop?  Check out some of these resources:

What are all the “-omes” in science?

If you’re into yoga, you may be very familiar with “OM” or if you’re an electrician with “ohms”, but in science, we use “-ome” in a very different way.  To start off, let’s give a silly example: the studentome (which I’m fairly sure does not actually exist).  This would be the study of all “students” in a certain place.  Maybe we’re interested in all of the students in a particular high school or college, and they can be categorized and better understood by looking at the distribution of their ages, their heights, their grade level, their clothing style, etc.  The studentome would be different in an inner city school compared to a private Catholic school, and understanding these differences could help to improve or change aspects of the studentome in a certain place. The study of the studentome, would be called studentomics.

So what does this “-ome” mean? In Greek: “-ome” means “all” or “complete”. So whenever scientists put “-ome” at the end of a word, they are talking about all of something (like with the “studentome” all of the students), and in biology this usually is referring to all of something in an organism or a particular cell type.  It’s not clear when scientists started using words ending in “-ome”, though the word biome was coined in the early 1900s.  The modern usage likely started in the early 1990s as technologies, like computers and DNA sequencing, allowed scientists to study all of something in an organism with more ease.  A derivative of the “-ome” is “-omics”, which is the study of all of something in an organism or a cell.  This terminology is so common, there is even a wiki dedicated to “-omes” and “-omics” here.

Let’s explore some of the common “-omes” (you can find a more comprehensive list of scientific “-omes” here).

  • Genome: The most famous “ome” is the genome. The genome refers to all of the DNA in an organism.  In humans, this includes all 23 pairs of chromosomes (number 1-22 and the two sex chromosome, XX if you are female and XY if you are male). Scientists study the genome to understand the genetic blueprint of DNA because DNA codes for proteins, which are the functional machines that do everything in a cell.
  • Transcriptome: For DNA to make a protein, the DNA needs to be “transcribed” in RNA first (read more details about this process here).  All of the RNA in a cell is called a transcriptome.  Scientists study the transcriptome because not all DNA is “turned on” to make proteins in every cell.  This helps explain why certain cells look different (skin cells look different than eye cells) and have a different function (skin cells provide a barrier from the environment and specialized eye cells allow you to see).
  • Proteome: And this brings us to the proteome, which is all of the proteins in a cell or organism.  Since proteins are what’s actually doing stuff in a cells, by understanding what proteins are present in certain cells, scientists are able to better understand how those cells function. And in the case when there are problems, for example in cancer cells, it can help understand why there is a problem and possible ways to fix it.

omes

  • Interactome: Proteins interact with one another in a variety of ways.  The interactome maps all of these interactions.  The interactome is also different in different cell types because the proteins expressed are different in different cell types, so there are many interactomes
  • Metabolome: Even though proteins are the machinery in a cell, there are lots of other small molecules and chemicals called metabolites.  For example, glucose is a metabolite that is broken down to produce energy.  All of the metabolites in an organism are called the Metabolome.

And there are hundreds more of these “-omes”!  This “omeome” (originally and jokingly coined here) has even seeped into more popular culture.  For example, the Facebookome, described as “The totality of facebook social network connections and nodes information such as people’s names, relationships, and multimedia contents.”  Although it may seem to be an unnecessary wordy trend, these “-omes” and “-omics” are necessary for scientists to better understand health and disease.