Exploring the miracle of life

Microbial dark matter

For a long time, microbiology worked exclusively on microorganisms that were first cultivated and multiplied in the laboratory and then isolated in order to be examined. The problem was that most microorganisms were lost during this propagation process and thus eluded analysis, a phenomenon called “microbial dark matter” in analogy to astronomy, denoting the millions of bacterial species that populate the world but which nobody knows about. When he worked on his doctoral thesis, Michael Wagner already developed new methods of identifying the microorganisms in the “dark matter” in terms of molecular biology and of specifically staining them without multiplying them in the laboratory. He also discovered completely new development strains and species. He remembers that period in the 1990s as “a time of pure wonder”. “Even in the most trivial of samples, on a shoe sole, for instance, we found new species and, for the first time, were able to specifically mark them with a fluorescent dye and observe them under the microscope.”

Watching bacteria feed

The scientist was not content simply to map the diversity of these microbes: he also wanted to find out what functions were fulfilled by the bacteria he detected. His guiding principle: “You are what you eat,” meaning he derives what the bacterium does from what it feeds on. Wagner uses isotope-labelled feed, which the bacteria absorb into the biomass when feeding. But how do you watch a creature a thousandth of a millimetre in size eat? He uses processes from materials science or surface chemistry for this purpose. The Centre for Microbiology and Environmental Systems Science boasts the only so-called “NanoSIMS” in Austria, a device worth three million euros which shoots an ion beam at cells and then uses mass spectrometry to determine whether cells are isotope-labelled or not. However, such investigations are very time-consuming, which is why only relatively few samples can be examined.

Revolution in microbiology

Wagner's research group was the first in the world to watch bacteria feed in their natural ecosystem. “That was a revolution,” he notes, “we were the first not only to map, but also to begin to understand the functions of microbes.” Another revolutionary step was the sorting of the bacteria according to their functions. In cooperation with Roman Stocker from ETH Zurich, the Vienna researchers have developed a finger-sized cell sorting machine that uses laser beam and Raman microscopy to remove isotope-labelled cells from a sample as though with tweezers, in order subsequently to cultivate them or to examine their genome sequence.

Intestinal bacteria and diseases

In recent years, microbial communities have also taken on increasing importance in human medicine. Today it is known that the composition and activity of intestinal bacteria plays an important role in the development of many diseases and that there is even strong evidence for the involvement of the intestinal microbiome in the development of certain types of cancer and mental disorders such as depression. Medical experts are therefore looking for ways of specifically modifying the intestinal microbiome, which is a more complex undertaking than one might think. Every human being has a specific composition of intestinal bacteria, which is not only influenced by factors such as genetics, nutrition, exercise or medication.

“The type of birth is already decisive - whether through the birth canal or by Caesarean – and whether a baby is breastfed or not,” says the biologist and adds: “This is why certain drugs have an effect on one patient but not on another, because bacteria convert and degrade active substances. If you host a bacterium that degrades the active ingredient of the drug, it cannot work or only to a lesser extent.” Targeted changes in the intestinal microbiome are not a trivial affair because each patient represents an ecosystem in its own right. Probiotics contain the same types of bacteria for all patients. “For many patients, taking these probiotics is like putting a cow and a bull in the rainforest, where they will not develop a population because they have too many enemies there,” is how Wagner, a great lover of nature, illustrates the complexity. He sees the future in personalized probiotics that are tailored to the intestinal microbiome of the particular patient.

Expertise for the Vienna General Hospital

Wagner sees human microbiome research as one of the most important research areas of this century. In the centre he heads, several working groups are focusing on this topic. In close cooperation with the Medical University of Vienna they are making their expertise available to the Vienna General Hospital as part of the “Joint Microbiome Facility” founded in 2018. “Human microbiome research uses concepts and methods of environmental microbiology, which has been investigating complex microbiomes intensively for decades. The greatest breakthroughs can be expected at the interface between environmental and medical microbiome research. We would like to make a contribution to ensuring that Austria does not miss the boat when it comes to developments in this area which is so important for public health, and we are grateful for the opportunity to set up such a facility here in one of the largest hospitals in Europe,” says the expert. Wagner is convinced that ten years from now, the hospital will not only a take a blood sample but also a stool sample for microbial analysis as a standard procedure.

Nitrogen cycle

One of Wagner's main research areas deals with microorganisms that play a central role in the global nitrogen cycle. The Earth's natural nitrogen cycle is massively influenced by human activity, above all by fertilizers used in agriculture. In the ground, the ammonium used for fertilization is first converted to toxic nitrite by microorganisms and then into the slightly less harmful nitrate.

Dead zones in the water

This two-stage process of “nitrification” is of tremendous ecological significance. As nitrate washes out of the soil with particular ease, more than half of the fertilizer is lost and leads to nitrogen pollution of groundwater, rivers, lakes and oceans. The eutrophication of bodies of water turns them into “dead zones”. “In the Gulf of Mexico, where large areas of the USA are drained by the Mississippi and the Atchafalaya, there are huge so-called dead zones where algae grow explosively as a result of over-fertilization. The algae are broken down by bacteria that deplete the oxygen in the process, which is why no higher life can survive there any longer,” notes Wagner in giving an example of these dynamics.

The problem of nitrous oxide

Nitrogen fertilization causes yet another problem: during nitrification, the microbes release “laughing gas” (nitrous oxide), a greenhouse gas that is 300 times more powerful than carbon dioxide and destroys the ozone layer. While these nitrifying microbes are very problematic in agriculture, they play an important role in water treatment plants by converting the ammonium from the wastewater into atmospheric nitrogen. For this process they join forces with so-called denitrifying bacteria. However, it is precisely under oxygen-poor conditions as they are found in sewage treatment plants and also in fertilized soils after rainfall, that a particularly large amount of nitrous oxide is released. For this reason, microbiologists have been looking for a long time for nitrifiers that release less nitrous oxide and have wondered whether it was possible to specifically promote their growth in such systems.

“Comammox” – a scientific breakthrough

In 2015, Wagner’s research group was exploring new nitrifiers when they achieved another scientific breakthrough in cooperation with Holger Daims from the University of Vienna: they discovered “Comammox” bacteria (complete ammonia oxidizers). These bacteria are particularly efficient nitrifying agents that convert ammonium into nitrate on their own without the need for microbial partners. And, as a great boon, only small amounts of nitrous oxide are released in the process. “A green nitrifier, if you like,” laughs Wagner. For the time being, the Viennese researchers are the only ones worldwide who are able to breed Comammox in the laboratory. They are now investigating the growth conditions of this bacterial species. Their vision for the future is to use these findings to promote the growth of Comammox in the environment in order to develop new, more environmentally friendly approaches in agriculture, wastewater treatment plants and drinking water treatment.

A formative childhood experience

For the 54-year-old Wagner, the decision to studying biology was a gut decision, as he had always been interested in the diversity of life. He had not planned ending up in microbiology, but perhaps this trajectory was fostered at the unconscious level by childhood experiences that had a great impact on him. When he was seven years old, his mother suddenly and unexpectedly fell ill with a brain infection caused by an unknown pathogen. Family life thus changed radically from one day to the next. “The question as to what illness my mother suffered from and how she could best be treated has been with us for many years,” recalls the father of four. “It is, perhaps, not entirely by chance if today I find myself developing methods for the identification and functional investigation of unknown microbes,” he speculates.

From a “village of indomitable Gauls” to world renown

After studying biology at Munich Technical University, the young postdoc spent a year of research at the US Northwestern University in Evanston and then returned to the Technical University of Munich and qualified as an associate professor. In 2003, he was called to Vienna and took on the position of Professor of Microbial Ecology at the University of Vienna. He was accompanied by Holger Daims, Matthias Horn and, a little later, Alexander Loy, formerly doctoral students in his team and, today, highly distinguished scientists. “We sat together in an office and planned the new department,” he remembers these early days. “We felt like the small village of indomitable Gauls: that forges a great sense of community.” One can still see this community spirit in the photo taken at the award ceremony for the Wittgenstein Prize 2019, where he is visibly buoyed by his colleagues’ cheers. “The others are well and truly delighted for me. That's the most beautiful thing.” Today, the “small Gallic village” boasts about 150 people from 29 countries. Fifteen of them are principal investigators and six of them have received one of the highly coveted ERC grants from the European Research Council.

Developing the technology further

For the next few years, Wagner anticipates that he will focus on further methodological improvements. Physicists at Boston University have developed a new Raman technology. This is a particularly fast type of chemical microscopy that reveals not the shape but the chemical composition of the cells. Wagner now wants to adapt this technology for microbial research in collaboration with the American teams. “This is high-risk research,” notes Wagner, because nobody has yet tried out whether this sophisticated technique can be combined with fluorescence microscopy and therefore be applied in the analysis of microbial communities. “If it works, we can watch the bacteria feeding almost in real time and see, for example, how different diets affect the activities of intestinal microbes.” That would represent yet another scientific breakthrough. Another thing becomes very clear: microbial research is strongly technology-driven. It is an area where technological progress often triggers the decisive next step.

More risk commitment

Wagner would like to see more commitment to risk from research funding institutions in general. He is very positive that the Austrian Science Fund FWF is moving in this direction with its newly conceived 1,000 Ideas Programme: “With this programme there is no need for any prior work before you apply for funding, which is very good, because there is currently a large funding gap in the area of high-risk research."