Wednesday, January 23, 2013

Culturomics- A Perspective


Fig 1: Core and variable components
of the human microbiome. Source 
     Give a moments thought to this. There are several set of organisms seen in association with humans as microbial flora. Much of the microbes have not been studied in detail, that is demanded by the science. Reason? its simply difficult to culture them in the laboratory. With the invent of DNA studies, a large number of bacteria have been studied with strategies such as the metagenomics. A detailed DNA analysis has also yielded insights into culturing bacteria's, that we are other wise unable to grow with common known set of laboratory conditions. In literature you would find a term "non-culturable bacteria". The term is often misleading to many students. The non cultivable conditions arise from our inability to grow them in artificial conditions rather than the microbes capability to grow. In the scientific writing it is often referred as VBNC (Viable but non-culturable) organisms.

    There is matter of interest here. Bacteria sure is capable of dividing (in other words reproducing). If i had to collect a few grams of urine sample from a normal person and plate it on all the available media, am probably going to end up with maybe 20 different types of organism. Or if am lucky enough, maybe a total of 50 species. But, guess what? There are at least another 1000 (probably one more zero), that is sitting in the sample not giving you a colony. But they have the capability to reproduce given the right conditions. That is the limit to Clinical Microbiology when it comes to culture. The effect is more greater when it comes to culturing bacteria from soil or other super microbe rich sources.

    Why am i boring you with the damn culture? After all we have PCR or DNA sequencing methods that can tell all the range of organism is there. Why culture? think about it for a minute and you have a set of answer. If you can grow the organism, you have to study and tease out its biology. DNA sequence and PCR (or other sophisticated methods to say) can tell you the gene part. As biologists, we are often interested in knowing the dynamics of the organism. And culture is a great way of doing it (Pls note, its not the only way).

    3 paragraphs of explanation to come to this one point of what am pointing towards. The post is all about "Culturomics". When I first heard this term, had the same reaction as you probably do. Yet another "Omics". But as I came to understand more and more what the idea seem to be, I begin to take a grasp of what exactly it is. But then am not a big fan of the "-omics" used for everything.

      Many a times crowning a title creates a confusion. Culturomics has two meanings. The first one is the field that focus on studies of linguistic and cultural phenomena (Link) and the second is one which focus on method of culturing bacteria (the focus is on improvising the culture technique).

    Culturomics is classically defined as "an approach allowing extensive assessment of the microbial composition by high-throughput culture". The definition is often seen ion contrast to metagenomics and possibly an alternative. An important digression here. Metagenomics is a sequencing approach (using pyrosequencing or 3rd generation sequencing methods) and aims to construct a map based on the DNA studies, whereas culturomics uses studies based on growth characters in artificial set of conditions.

Fig 2: Identification of bacteria in the human gut
by culturomics and metagenomics.
    Alright. here's the paper that came first on the field that's worth a bit of discussion. This paper is by Lagier etal; In this study, stool samples from 3 people (2 lean African and one obese European), were analysed by 212 different culture conditions of culture. The study recovered 32500 colonies which was traced to 340 species of bacteria from seven phyla and 117 genera, five fungi, and a giant virus (Senegalvirus). Its not the recovery itself that am impressed about. Its the fact that 212 variety of conditions that then lead to recovery. A parallel meta-genomic approach was used for comparison, yielded 698 phylotypes, including 282 known species. By using a set of conditions they could culture out nearly half of the population that exists. That's mind blowing. So if you could get more conditions, probably they would get more. But for a start, it is massive step. Just make a note here that the metagenomics identifies probably about 50-60% of what is actually present. The Fig 2 from paper says it all.

    So what was the novel culture conditions that was brought in? It was use of rumen fluid and sterile fresh human stools. Selectivity in culture was tried by removal of E. coli by specific lytic phages, various antibiotic mixtures, heat destruction of non-sporulating bacteria, exotic range of selective media, amoebal co-culture. The selectivity gave a chance for other bacteria's that are more slow in growing (I call the shy growers) to get a hold and be cultured. But I could make a more general point here. The paper had concentrated more on culturing conditions very similar to native growing conditions. That makes sense. Perhaps we microbiologists should try to culture the microbe "in their conditions" rather than "our conditions".

     A very fine explanation on the use of culturomics and its contrast to metagenomics is published recently by G. Greub. The paper provides compelling evidence towards the new hot topic "Culturomics". But, I gave it some thought. We have been cultivating a variety of microbes in the lab. Though, Microbiologists have not successfully cultivated all the organisms, they aren't that bad too. Lets assume that we have not been able to grow a bacteria X, but happened to sequence its genome. And by bioinformatic approach, we concluded that the X is missing a pathway for making some important nutrient Y. Under normal culture conditions, the Y is not provided in the growth conditions. But if you substitute it, you now grow the bacteria X. This is not my naive approach. This method has been used for a longtime now, to culture organisms that were otherwise thought to be Non culturable. But they never gave this approach a entire field name, for it doesn't sufficiently represent a territory in itself (the way genomics, Proteomics etc do), and am little hesitant to call it with a suffix- "omics".

    But, perhaps right now this is a naive field and just begun to evolve. But whatever is the name is the take home excitement is that, we are now thinking of the classical microbiology culture once again, and trying to figure out how to grow the ones that are otherwise "shy growers". And an important point, this is a "Select in-out approach".
Lagier JC, Armougom F, Million M, Hugon P, Pagnier I, Robert C, Bittar F, Fournous G, Gimenez G, Maraninchi M, Trape JF, Koonin EV, La Scola B, & Raoult D (2012). Microbial culturomics: paradigm shift in the human gut microbiome study. Clinical microbiology and infection : the official publication of the European Society of Clinical Microbiology and Infectious Diseases, 18 (12), 1185-93 PMID: 23033984

Greub, G. (2012). Culturomics: a new approach to study the human microbiome Clinical Microbiology and Infection DOI: 10.1111/1469-0691.12032

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