Gearing Up for Microbiome Research

George Weinstock, PhD, professor and director of microbial genomics at the Jackson Laboratory for Genomic Medicine, talks to contributing editor Tanuja Koppal, PhD, about the differences between microbiology and microbiome research. He talks about the big challenges facing the microbiome field and discusses ways in which lab managers can better equip their labs and build expertise to face those challenges.

Q: How does microbiome research differ from traditional microbiology?

A: Microbiology pays attention to individual species of a microorganism; for instance, studying species of bacteria like Salmonella or Escherichia coli. When doing a microbiome study, you are looking at many different species and microorganisms at the same time. It involves studying the community of microorganisms as a whole. It’s not as easy as studying a single species, where you can isolate, purify, and culture the cells. Many of the species found in a microbiome may have never been grown in a culture. Traditional microbiology does have branches like environmental microbiology or microbial ecology where you study communities of microorganisms, but that is now amplified and expanded by microbiome research.

Q: How do the laboratories for microbiology and microbiome research differ in terms of their activities and setup?

A: Many of the microorganisms found in a community have never been cultured; hence, microbiome labs are driven by methods that are usually culture- independent. The most important of those methods is DNA sequencing. In a microbiome study, a nucleic acid preparation is done to isolate the DNA from the entire sample, which can be a saliva, nasal, or stool sample. The sequencing is not done on individual DNA separated from the various species seen in the sample. Then a bioinformatics analysis is done on the sequences obtained from the mixture to try to figure out what organisms may be present. So you gather a list of all the different organisms present in the mixture without trying to culture each of them separately. In microbiology you are often purifying and culturing species and trying to identify them separately. In microbiome research you rarely do that. Instead, you rely on DNA sequencing and the associated bioinformatics tools to convert mixtures of DNA sequences into a list of organisms that may be present.

Q: What types of microorganisms are typically studied?

A: The predominant microorganism in the human body is bacteria, and bacteria is mainly located in the intestinal tract. There are also fungi and eukaryotic microorganisms that are found in abundance in many different body sites. Fungi, for instance, are more abundant in the mouth and skin than in the gut. Most healthy people also carry viruses without showing any sign of disease or infection and, hence, viruses are also considered to be a part of the human microbiome. There are various types of DNA sequencing that need to be done in order to identify all these different types of microorganisms. For bacteria, you sequence the 16S ribosomal RNA gene. Similarly for fungi and eukaryotes, you sequence the 18S ribosomal gene or the internal transcribed spacer (ITS) region. Viruses do not have ribosomal RNA, so one has to do shotgun sequencing to look at the entire nucleic acid preparation of the sample and go through all the sequences to see which of them are viral in origin. Viruses can have both, DNA or RNA, so you have to make an RNA preparation in addition to the DNA prep, and then convert the RNA into cDNA in order to sequence it. Thus, in order to get an exhaustive view of the microbiome, one has to do many different types of sequencing— targeted sequencing of 16S, 18S, or the ITS region; shotgun sequencing; RNA sequencing; and much more.

Q: How can traditional microbiology labs start building an expertise in genomics and bioinformatics?

A: There are options for outsourcing some of the work to companies that already have the expertise to do that. Depending on the type of sample and how much you need to oversee the process, you can either send out the samples—or the DNA preparation from those samples—to outside labs.

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Q: What are some of the big challenges in conducting microbiome research?

A: Generating the data using DNA sequencing is pretty straightforward. The parts that introduce variability and bias in the results are often associated with how the samples are handled initially. In microbiology there is some amount of tolerance associated with sample handling since, very often, you only want to know if a certain species exists in the sample. In microbiome research, you want to know the types of organisms present in the microbial community along with their relative abundances. Since this work is quantitative, the handling of the samples becomes very critical. With some organisms being hardier than others are, the whole structure of the community can be affected during handling. Some samples, such as stool, saliva, or nasal samples, can be collected and frozen right away, but in some situations when that’s not possible, the preservation and handling can affect the microbiome structure, and hence the results. Sample handling is always going to be a big challenge because you are looking at community structures and you want to preserve their relative stoichiometry. At the other end of the process, once you have a list of the organisms present in the community and their relative abundances, comparing different communities across samples becomes a challenge too. The statistical methods that are necessary to draw really strong conclusions get complicated and more advanced. That downstream analysis of the complex microbiome data becomes very tricky.

Q: What types of microbial work are you involved in?

A: The two things we are most interested in are comparing the microbiomes of different sets of people—healthy people versus those having a particular disease, old versus young people, or pregnant versus nonpregnant women, and others—to look at changes in the microbiome in different tissues. Once we identify the differences in the microbiomes, then we try to identify the species that have changed and whether their abundance has gone up or down. If there are some organisms found that have never been cultured, then we try to culture them. The idea is to identify the mechanism of diseases by understanding what specific species have changed in the microbiome and then to try to figure out how those changes have contributed to the disease. We study lots of different diseases and work with clinicians and hospitals to provide us with samples, and they work with us to study the mechanistic role of the microbiome in disease. The other area we are working on is geared toward improving the diagnosis of infectious diseases. Sometimes diagnostic labs at hospitals are only looking to identify common pathogens and are not able to identify the less-common organism that may be responsible for the infection. This is particularly true for patients who are immuno-suppressed, as they can get infected with pathogens that would not regularly infect a healthy individual. We would like to do a microbiome analysis of all those patients to figure out what microorganisms are present in them. We are working to make such general types of pathogen diagnosis faster, cheaper, and to be able to do this with a higher throughput.

Q: Will microbiome studies have an impact only on life sciences and medical applications?

A: Microbiome research is certainly ongoing everywhere. In agriculture, the animal microbiome can affect animal health and agricultural production. Plants have microbiomes too; they have an interesting microbiome around their roots, which is essentially like an inside-out human intestinal system. That’s where nutrients are absorbed and waste products are released, and that microbiome interacts with the soil as well as with the plant. Microorganisms in the environment have been around far longer than plants and animals, and they have had a big impact on the geology and the processes taking place on Earth. There is an Earth Microbiome Project that looks at microbiomes present in different habitats. So microbiome research is a huge field that goes well beyond human health. There is a trend to set up microbiome centers in universities that will serve as more than just core facilities. These university-based microbiome centers bring together faculty who are involved in various types of microbiome research. There is a rich set of resources out there for anyone looking to study microbiomes to see how it’s done.

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