Last Updated on September 17, 2016 by Patricia Carter

SUMMARY:  This genius 2 week time-lapse video by Harvard Medical School lab lets you watch bacteria become antibiotic resistant.  This experiment is thought to be the first large-scale glimpse of the maneuvers of bacteria as they become superbugs as they encounter increasingly higher doses of antibiotic and adapt to survive — and thrive — in them!  Also read what to do if prescribed an antibiotic. Why? Antibiotics severely affect microbial diversity in the gut for months after exposure (4 months for clindamycin, 12 months for ciprofoxacin)…  An added bonus of this experiment is the rewriting of lab research tools:  The use of a large scale petri plate now opens up new ways for researchers to think about and conduct novel experiments!

For most folks, antibiotic resistance is conceptual and abstract.  Most don’t understand why there now is a push for judicious use of antibiotics.  For those details, see WHO, World Health Organization Vows To Combat Drug Resistance, with more here, and here for WHO pdf.  

“Getting more people to understand how quickly bacteria evolve antibiotic resistance might help people understand why they shouldn’t be prescribed antibiotics.  The drug resistance is not some abstract threat. It’s real.” —Tami Lieberman, member of Alm Lab, an evolutionary microbiologist at MIT.  As quoted in WATCH: Bacteria Invade Antibiotics And Transform Into Superbugs.

In humans, antibiotics severely affect microbial diversity in the gut for months after exposure and enrich genes associated with antibiotic resistance,  One course of antibiotics disrupts gut microbiome for a year, 2015, explains:

•  Microbiome diversity in feces was significantly reduced for up to 4 months for clindamycin and up to 12 months for ciprofloxacin.  Contrast this to microorganisms in the oral cavity and saliva which showed signs of recovery in as little as a few weeks after drug exposure.
• Amoxicillin had no significant effect on microbiome diversity in either the gut or oral cavity, but it was associated with the greatest number of antibiotic-resistant genes.
• Researchers saw a decline in the abundance of health-associated species that produce butyrate, a substance that inhibits inflammation, cancer formation and stress in the gut.

“Certainly we cannot live or survive without antibiotics; that’s out of the question. But there are situations when we should not use them, like when there are no evidence-based reasons.”

 —One course of antibiotics disrupts gut microbiome for a year, 2015,

Practically, be vigilant and advocate for yourself when in the situation where antibiotics are being prescribed.  

Ask for delay on starting the antibiotic until the culture results are in.  In the very least, make sure the least broad spectrum antibiotic possible is prescribed.  Many doctors will now agree.  

 Also included in this post is guidance on what to do if you have to take an antibiotic and Dr. Rob Knight ‘s Eleven Point Punch List of things that seem to be beneficial to the microbiome as learned from the American Gut Project data. 

The Harvard experiment allows you to see with the naked eye  the WHY behind judicious use of antibiotics.  

In the video, evolution appears to march across an enormous black petri dish (2 foot by 4 foot) that is plated with vertical bands of successively higher doses of antibiotic.  Watch evolution occur as bacteria move, adapt and mutate, die, and survive.  The bacteria Escherichia coli (e-coli) built resistance to a 1,000-fold to the initial antibiotic  trimethoprim dose.  Another antibiotic tested, ciprofloxacin, built resistance to 100,000-fold to the initial antibiotic dose.     

The Evolution of Bacteria on a “Mega-Plate” Petri Dish

“We can see that by this process of accumulating successive mutations, that bacteria which are normally sensitive to an antibiotic, can evolve resistance to extremely high concentrations in a short period of time.”

The large petri dish is called Microbial Evolution, and Growth Arena (MEGA) plate.

Scientists from Harvard Medical School and Technion-Israel Institute of Technology designed the simple experiment to observe how bacteria move as they become impervious to drugs.  The large 4-by-2-foot spatial petri dish was built by postdoc Michael Baym at Harvard Medical School, Roy Kishon’s lab.  The movie was compiled from time-lapse imagery every 10 minutes for 11.7 days, and played at 30fps (18000X speed). Each second of video is approximately five hours of real time.

MEGA Plate Description.  The outermost rims of the dish contained no drug. The next section contained a small amount of antibiotic — just above the minimum needed to kill the bacteria — and each subsequent section represented a 10-fold increase in dose, with the center of the dish containing 1,000 times as much antibiotic as the area with the lowest dose.

Pearls to note while watching the time-lapse video of bacteria becoming antibiotic resistant.

The white bacterial colony creeps across the MEGA plate allowing visual observation of bacteria mutation and selection:

Bacteria (white) grow up to the boundary where they can no longer survive.  The colony pauses when it hits the first band of antibiotic, creating a stark border between the white colony and the black petri dish.  Mutants, capable of surviving the higher concentration of antibiotic appear and invade the new band.  More dots appear and they start growing, racing to the next, stronger band of antibiotic. The bacteria are evolving.  Subsequent steps require further mutations.  After about 11 days, resistance to over 1000 times as much antibiotic as was originally toxic evolves.  After almost two weeks of real time have passed, they’ve become resistant to the strongest completely taken over the kitchen-table-sized petri dish. —The Evolution of Bacteria on a “Mega-Plate” Petri Dish and WATCH: Bacteria Invade Antibiotics And Transform Into Superbugs

Key insights for bacteria behaviour from the MEGA Plate

From Harvard Gazette, A cinematic approach to drug resistance, Sept. 8, 2016:

• Bacteria spread until they reached a concentration (antibiotic dose) in which they could no longer grow.
• At each concentration level, a small group of bacteria adapted and survived. Resistance occurred through the successive accumulation of genetic changes. As drug-resistant mutants arose, their descendants migrated to areas of higher antibiotic concentration. Multiple lineages of mutants competed for the same space. The winning strains progressed to the area with the higher drug dose, until they reached a drug concentration at which they could not survive.
• Progressing sequentially through increasingly higher doses of antibiotic, low-resistance mutants gave rise to moderately resistant mutants, eventually spawning highly resistant strains able to fend off the highest doses of antibiotic.
• Ultimately, in a dramatic demonstration of acquired drug resistance, bacteria spread to the highest drug concentration. In the span of 10 days, bacteria produced mutant strains capable of surviving a dose of the antibiotic trimethoprim 1,000 times higher than the one that killed their progenitors. When researchers used another antibiotic — ciprofloxacin — bacteria developed 100,000-fold resistance to the initial dose.
• Initial mutations led to slower growth — a finding that suggests bacteria adapting to the antibiotic aren’t able to grow at optimal speed while developing mutations. Once fully resistant, such bacteria regained normal growth rates.
• The fittest, most resistant mutants were not always the fastest. They sometimes stayed behind weaker strains that braved the frontlines of higher antibiotic doses.  
• The classic assumption has been that mutants that survive the highest concentration are the most resistant, but the team’s observations suggest otherwise.

“What we saw suggests that evolution is not always led by the most resistant mutants.  Sometimes it favors the first to get there. The strongest mutants are, in fact, often moving behind more vulnerable strains. Who gets there first may be predicated on proximity rather than mutation strength.”  

— Michael Baym (first author on study) research fellow in Systems Biology, Kishony Lab, Harvard Medical School, A cinematic approach to drug resistance, Sept. 8, 2016.

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MEGA plate concept opens up new research opportunities in lab

Getting away from the small, classic petri dish opens up thinking outside the box for a lot of research.  The giant petri dish opens up a spatial dimension that has been missing the lab.  Up to now, research uses tiny plates containing homogeneously mixed doses of drugs.  For comparison, the MEGA was filled with 14 liters of agar, a seaweed-derived jellylike substance commonly used in labs to nourish organisms as they grow. 

The giant MEGA petri dish better mimics real-world bacteria exposure since for bacterial evolution, space, size, and geography matter.  Different challenges are faced for the bacteria as they move across varying antibiotic strength environments.  The MEGA dish does not  perfectly mirror how bacteria adapt and thrive in the real world and in hospital settings but it moves closer to real world environments than the traditional lab small homogeneously mixed petri plates.  A cinematic approach to drug resistance, Sept. 8, 2016.

If you can see it, maybe you can start to study it.

“Hopefully this will put back in people’s minds how important the spatial element can be.”  —Pamela Yeh, a microbiologist at UCLA who was not involved in the experiment.

From the study, Journal Science, Spatiotemporal microbial evolution on antibiotic landscapes, Sept 9, 2016:

“… [the MEGA plate] allowed visual observation of mutation and selection in a migrating bacterial front. While resistance increased consistently, multiple coexisting lineages diversified both phenotypically  and genotypically. Analyzing mutants at and behind the propagating front, we found that evolution is not always led by the most resistant mutants; highly resistant mutants may be trapped behind more sensitive lineages. The MEGA-plate provides a versatile platform for studying microbial adaption and directly visualizing evolutionary dynamics.”

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Some posts to read concerning  WHAT to do if you have to take an antibiotic are:

1. What to do if taking ANTIBIOTICS, MICROBIOME —  Diet hugely impacts microbiome.  This post details strategies that especially support and build up the microbiome, Florastor,  plus info on why avoiding the acid suppressors is important. 

   

   Antibiotics severely affect microbial diversity in the gut for months after exposure and enrich genes associated with antibiotic resistance.  Follow the suggestions in #1 to preclude pathogenic bacteria from gaining traction in the antibiotic nuked microbiome.    One course of antibiotics disrupts gut microbiome for a year, 2015, explains:

    Microbiome diversity in feces was significantly reduced for up to 4 months for clindamycin and up to 12 months for ciprofloxacin.  

   Amoxicillin had no significant effect on microbiome diversity in either the gut or oral cavity, but it was associated with the greatest number of antibiotic-resistant genes.

   Researchers saw a decline in the abundance of health-associated species that produce butyrate, a substance that inhibits inflammation, cancer formation and stress in the gut.

   “Certainly we cannot live or survive without antibiotics; that’s out of the question. But there are situations when we should not use them, like when there are no evidence-based reasons.”

    —One course of antibiotics disrupts gut microbiome for a year, 2015,

   Practically, be vigilant and advocate for yourself when in the situation where antibiotics are being prescribed.  

   Ask for delay on starting the antibiotic until the culture results are in.  In the very least, make sure the least broad spectrum antibiotic possible is prescribed.  Many doctors will now agree.  

2. US Antibiotic Resistance and WHO. Check out the slides in this post to see your states antibiotic use and risk of antibiotic resistance.
3. The post, PRESERVE & RESTORE LOSS OF MICROBIOME DIVERSITY IS AGGRESSIVE PREVENTATIVE MEDICINE, is perhaps the most reasonable mechanism to prevent antibiotic need in the first place, especially given that the average American has lost:

• 1/3 of their microbiome diversity, as estimated by Dr. Maria Gloria Dominguez-Bello as seen on this interview for the film “Microbirth.”  
• Another study estimates 1 in 4 have 40% less bacteria (see This Danish study, or read the scientific article here).  

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Dr. Rob Knight’s Eleven Point Punch List of things that seem to be beneficial to the microbiome:

One of the best ways to protect your health is to protect the health of your gut microbiome.  WE ARE SICKER WHEN WE HAVE LESS BACTERIAL Abundance and Diversity WITHIN OUR MICROBIOME.

Diversity in your gut microbiome is key to health.  

A diverse diet means diverse microbes which is key to diversity in your microbiome. 

This post summarized a lecture by Dr. Rob Knight wherein he summarized the American Gut Project data and gave this Eleven Point Punch List of things that seem to be beneficial to the microbiome:

1. Eat lots of  plants:  5 to 30 different varieties each week preferably.  See MY NOTES below for more explanation.  
2. Aging increases microbiome diversity:  Microbiomes are more diverse at age 50 to 60 then populations in their twenties  (see below slides).
   

   
3. Having an IBD diagnosis means your microbiome is altered.  NOTE:  Many chronic and autoimmune diseases are also following suit. Click the drop down menu on the right sidebar to see the microbiome — disease link.
4. The time of year alters the microbiome with a more diverse microbiome being with sun and outdoor exposure.
5. Antibiotics wipe the microbiome with some folks recovering relatively soon whereas others do not recover the pre-antibiotic microbiome even one year later.
6. Males vs females:  The sex for a  given microbiome can now be accurately predicted.
7. Sleep 8 hours for a more diverse microbiome.  Less than 6 hours yields a less diverse microbiome.
8. BMI but it only subtly affects the microbiome.
9. Plants: eating 6 to 10 each week is good, but eating 30 plus different varieties is best.  (See further discussion below.)
10. Alcohol: one drink of red wine is helpful, more than one reduces diversity.  Red wine has over 100 different polyphenols, providing a feast for your microbes.  Cheers!
11. Frequent exercisers have a more diverse microbiome and it is best if exercise is outdoors rather than indoors.

Eat lots of  plants: 5 to 30 different varieties each week preferably  

This finding is so profound that Dr. Knight noted that “American Gut” will soon change participant food journal requirements to only ask for frequency on consumption of holistic food within the past month, instead of the three week food journal.  This change is warranted since the long term diet, especially meat and fiber consumption, has been shown to have the largest effect on the microbiome.

How Bad for you is a Boring Diet?  You may think you eat a varied diet, but our hunter-gatherer ancestors’ diet was actually twenty times more diverse. 

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One example of a bacteria becoming increasingly antibiotic resistant, CDiff.

Dr. Dale N. Gerding, an infectious diseases specialist at Loyola University Chicago:

• C. diff. is found in soil and water, even chlorinated water, and is a low-level contaminant in food. Most of us ingest C. diff. every day. In most people the myriad micro-organisms that normally reside in the gut protect against C. diff. infection.
• Antibiotics disrupt the healthy balance of micro-organisms. Freed of competition, C. diff spores can germinate and reproduce unchecked, and not only in people with compromised immune systems.  I can’t reiterate this enough:  PRESERVE & RESTORE LOSS OF MICROBIOME DIVERSITY IS AGGRESSIVE PREVENTATIVE MEDICINE 

Ampicillin, amoxicillin, cephalosporins, clindamycin and fluoroquinolones are the antibiotics that are most frequently associated with the disease [CDiff], but almost all antibiotics have been associated with infection.” –Wide Use of Antibiotics Allows C. Diff to Flourish

Even after infected individuals recover, about 5 percent continue to harbor the toxic strain in their stool for six months, and if they take another antibiotic during that time, the illness can recur.” —Maja Rupnik wrote in The New England Journal of Medicine

CDiff spores are resistant to heat, acid and antibiotics.  

They can be washed away with soap and water but are NOT inhibited by the alcohol-based hand sanitizers now widely used in health facilities.  Just use soap and water!!! –Wide Use of Antibiotics Allows C. Diff to Flourish.  

In conclusion, I hope you learned more about the importance of keeping your microbiome health strong to avoid antibiotics in the first place.  But if they are needed, this post should help give insights into things to do to help mitigate the microbiome nuke.  I also hope Harvard’s video leaves you with better appreciation of the real threat of bacteria increasingly becoming antibiotic resistant.

Last updated: September 17, 2016 at 12:26 pm  to add quote from video:  “We can see that by this process of accumulating successive mutations, that bacteria which are normally sensitive to an antibiotic, can evolve resistance to extremely high concentrations in a short period of time” and for SEO optimization by adding in Summary section, “Also read what to do if prescribed an antibiotic. Why? Antibiotics severely affect microbial diversity in the gut for months after exposure (4 months for clindamycin, 12 months for ciprofoxacin)…”  Prior update added following reference to the post, What to do if taking ANTIBIOTICS, MICROBIOME:  “Diet hugely impacts microbiome.  This post details strategies that especially support and build up the microbiome, Florastor,  plus info on why avoiding the acid suppressors is important.” 

Best in health thru awareness,