Intriguing impurities

May 23, 2013

Many microbiologists begin their workday by opening up the incubator shaker and taking out a culture flask; they then swirl it around to examine the progression of cell growth. They can examine the turbidity of the broth to tell if cells are reproducing as planned. Opening up the lid and sniffing the culture will tell seasoned microbiologists if they’re growing what they think they’re growing, because many bacterial species produce signature aromas. But what microbiologists cannot see are minor contaminants lurking within: a nightmare for controlled studies. 

Shrestha et al. recently flushed a contaminant from their culture flask. They were working with a particular strain of Geobacter sulfurreducens, strain DL1. Their main interest was in using experimental evolution to improve this strain’s natural ability to conduct an electrical charge. In their experimental setup, G. sulfurreducens DL1 would take electrons from acetate and donate them to a graphite stick anode with a potential of -400 mV, producing electrical current. They inoculated the device and let it incubate for five months, with regular additions of fresh medium. The outcome? At the end of the experiment, the “evolved” strain was now better at producing electrical currents, and was better able to form biofilms on the graphite anode.  

The next step was sequencing the genome of the evolved strain, referred to as KN400. The researchers discovered that the genome of the KN400 evolved strain was drastically different from that of the inoculated strain DL1. Over 27,000 single nucleotides differed between the two strains’ genomes, and KN400 had over 100 genes not found in DL1. Given these genomic differences, the researchers estimated that these strains must have diverged at least 1,000 years ago, and that such extensive divergence was unlikely to occur during the five months of incubation with a graphite anode.

At first they were stumped. The DL1 cultures they were working with had been meticulously carried though the microbiological ritual of single colony re-streaking for strain purity. They also used the same culturing techniques for DNA isolation and complete genome sequencing, from which they found no sign of the contaminating KN400 (even with an average of 80-fold coverage of each nucleotide of DL1’s genome). Where did KN400 come from? At this point it may have been easy to blame one of the overworked graduate students or postdocs who performed the experiments. Luckily, these researchers investigated further.

It turns out that KN400 was always lurking in their cultures, at a very low level. By amplifying and sequencing millions of copies of one gene, they were able to detect KN400 in the DL1 inoculum at about 1/1x105 copies. This drastically low abundance was quite stable, retained through single colony re-streaking. Inoculating the two strains at equal densities always resulted in KN400 decreasing to this frequency, and the only way to lure KN400 out of hiding was to incubate the mixed culture in the experimental evolution conditions of -400 mV, an environment where they later learned DL1 died off within two transfers.  Eventually the researchers were able to obtain a pure culture of DL1, by repeatedly diluting liquid cultures.

The implications for their research are not that profound. They still have a strain that can conduct electricity very efficiently, and now they have rid their DL1 culture of its contaminant. But what about the implications for microbiology? Low-level contamination is probably much more common that we think. And this study has shown that whole genome sequencing doesn’t always uncover those contaminants lurking within. The important question is whether these contaminants contribute to the phenotypic properties of “pure” cultures. The answer to this question has far-reaching implications for microbiology, past and present.

 

Picoplankton post

March 29, 2013
My first contribution to the Small Things Considered blog: 
 

A Day in the Life: Eavesdropping on Marine Picoplankton


Continue reading...
 

Mite-y resistance

January 16, 2013
When faced with a challenge, animals have the ability to physically move away. Plants are not so lucky. Their interactions with the soil prevent them from up-rooting and finding a better place. But this does not mean that plants don't fight back. Indeed, plants produce an astonishing diversity of chemicals to ward off invaders, such as the ubiquitous arthropod pests. Many of these naturally produced chemicals are analogous to manufactured pesticides, in that they provide a strong selective pr...
Continue reading...
 

RNA for the bees

December 31, 2012
Becoming a beekeeper can be overwhelming at times. One of the most difficult things for me is figuring out how and when to treat the hives to prevent pathogens and parasites from becoming too numerous. We have attended an Integrated Pest Management course but only walked away with a list of offenders and their chemical treatments, which we had already learned through online research. Why not just treat for all, as often as possible? I'd rather not dump these chemicals (several of which are an...
Continue reading...
 

Reveling in Ronin

December 21, 2012
The Ronin Institute is a group of scholars that are trying something different.  Recognizing that one need not work at a university to contribute to knowledge, scholars of the Ronin Institute will be conducting research independently, as they find time in between life's activities (e.g., kids, hobbies, other jobs).  This is a timely endeavor as many more PhDs are being obtained than faculty positions are available.  And for many of us, university jobs are becoming less attractive as hours inc...
Continue reading...
 

Probiotic puzzle

December 4, 2012
Probiotics are live bacteria, formulas of single or multiple species, provided to help establish a healthy gastrointestinal tract.  Probiotics are a hot topic these days, and a potent marketing tool. Despite their popularity, we often don't know if they really work to relieve intestinal problems.  And for those probiotics that do work, we almost never know their mechanism(s) of action.   

Olier and colleagues have taken us a step closer to understanding the mechanism of probiotic action.  They...

Continue reading...
 

Bacterial relocation II

November 26, 2012
Continuing on the subject of bacteriotherapy, there was a fascinating paper out last month in PLoS Pathogens by Lawley et al. As I said in the last post, C. difficile infection of the gut is bad news, and one of the best cures is fecal transplantation.  Lawley et al. describe experiments in a mouse model that closely approximates the situation in humans.  They showed that the epidemic strain of C. difficile 027/BI is much more of a terror than other strains, as the bug was highly contagious a...
Continue reading...
 

Bacterial relocation

November 4, 2012
Warning: Here comes the icky diarrhea blog.  Because I primarily write about microbiota, this was only a matter of time as the literature is replete with gut microbiota studies.  This abundance is due to lots of money being poured into figuring out the role of the gut microbiota; they are likely to influence a plethora of diseases through their role in immune system development, and fecal samples are easy to obtain!

Fecal samples are also relatively easy to transplant.  This may sound gross be...
Continue reading...
 

Hurricane highway

October 31, 2012
Monday we made sure all of the objects in our yard were secure, to prevent the winds of Sandy from giving them flight.  We didn't know what to expect and didn't want to see our patio table jump through our window.  Luckily the winds didn't do any damage and we're all okay (and dry).

This got me thinking about bacterial dispersal, and how high winds like those in a hurricane really stir things up. Ocean water columns are mixed, ocean waters come ashore, terrestrial soil becomes airborne, tree l...

Continue reading...
 

Phages as fighters

October 16, 2012

 
Not all ingested bacteria will be able to colonize the intestines upon arrival.  Some bacteria may not be adapted to living in the GI tract, and are merely passersby waiting to be ejected into the environment to find their favored niche.  Other bacteria are happy to be in the GI tract where nutrients are abundant, but must contend with the other hungry microbes and the vigilant immune system.  When competition is fierce, a particular bacterial species may “win” by producing toxins that k...

Continue reading...
 
blog comments powered by Disqus

About Me


I am a biologist and writer. My most recent adventure has been to leave the comforts of academic life in the city, and start my own freelance business on a farm! At this blog you can read about all of this: from my thoughts and opinions on scientific research, to my trials and errors as a budding gardener and beekeeper.