Leaf to Field Newsletter, Issue 1: Feature Article

Instruments that make the invisible, visible

It is almost 38ºC. Inside the wheat crop, a woman with a broad-brimmed hat is looking intensely at a box with an arm that seems to be biting a wheat leaf. The box is measuring the exchange of gases during photosynthesis and this scene, repeated in numerous field experiments and laboratories around the world, produces data that is essential to the quest of improving yields of major food crops.

Improving photosynthesis is now recognised as one of the most effective ways to increase crop yields and technologies such as gas exchange measurement, have a pivotal role in achieving this. For the research undertaken within the Centre, this technology is a crucial link between researchers, projects and scales.

The invention of gas exchange instruments has allowed scientists to deepen their understanding of the photosynthetic process and its limitations. Photosynthesis researchers use these instruments constantly; travel to remote locations with them, find solutions to their leaks and problems, and everyday try to understand what they are saying to them about the gases that go in and out of their entrails.

 

 

“Gas exchange machines allow us to measure something that we cannot see and that happens within the cells of a leaf, which is quite amazing.”
Barbara George-Jaeggli.

 

 

Gas Exchange Instruments Profile
To measure gas exchange, scientists use a diverse range of instruments, from very portable automated machines to instruments that measure gas exchange at the canopy and atmospheric levels.

The term also covers instruments used in quite a wide range of science fields, from plant physiology to hydrology and chemistry.
The most common types currently used in plant biology clamp the leaf within the machine, creating a chamber which makes it possible to measure changes in Carbon dioxide (CO₂) and water vapour concentration in the air as it passes over the leaf.

Scientists can control environmental variables which affect photosynthesis, and can see the changes instantly graphed on a screen.

An important component of the technology are infra-red gas analysers (IRGA).These measure the specific changes in absorption of infra-red light due to CO₂ and water vapour.

 

When gases pass through plants

Gas exchange instruments measure how efficient a particular plant is at photosynthesising by estimating the exchange of CO₂, and water in the leaves. As Murray Badger, Director of the Centre points out “the properties of CO₂ and H₂O which relate to their absorption of heat or infra-red energy is what enables the whole technology to work. Without the heat absorbing properties of the gases we would not be able to detect them”.

Invented more than half a century ago, these instruments have since contributed to a vast number of new questions and answers on plant biology. A remarkable example, is the development in 1981, of the equations that are embedded in the instruments that make calculations of gas exchange by Susanne von Caemmerer, Deputy Director of the Centre, and Graham Farquhar, one of the Centre’s Chief Investigators. They developed a model of C3 photosynthesis to analyse gas exchange results and this model is now used worldwide.

Finely tuned, highly efficient, valuable, reliable and powerful are some of the words used by scientists to describe the modern versions of these instruments. “They offer valuable information about plant performance based on simple measurements”, says Dr Oula Ghannoum, a Chief Investigator within the Centre.

In a similar way to doctors measuring blood pressure and temperature of a patient to make a diagnosis on their health, plant scientists use gas exchange measurements, to be able to know what plants are actually doing. “Using gas exchange instruments is like using a stethoscope, in the sense that your measurements are in vivo and depend of the state of the individual at the moment you examine them,” says Viridiana Silva-Perez, a Centre PhD student.

These non-invasive instruments can measure variables without causing damage to the plants and the responses are instantaneous, showing changes in photosynthetic rate changes as they are happening.

An obvious advantage of the modern gas exchange machines is that they are extremely portable, which is indispensable for studies that take place in the field. This characteristic, has made them frequent travellers to every corner of the planet, from Tenant Creek (in the Northern Territory of Australia), to Europe, and from mangroves in North Queensland to boreal forests in Canada. “I took gas exchange instruments up a ladder to measure leaves on soil-rooted trees growing in whole tree chambers, and up a cherry picker in Oak Ridge, USA. I travelled with them to FACE [Free air Carbon Dioxide Enrichment] experiments around the world, including tropical pastures in Northern Queensland, temperate pastures in Palmerston North, New Zealand and a rice paddy in Morioka, Japan,” says Ghannoum.

 

 

Joining the dots

Perhaps the most impressive fact about gas exchange instruments is that they are used to answer scientific questions on hugely different scales, from the molecular level of the plant, to canopy and whole-tree gas exchange with the atmosphere. “They link different studies and scales from biochemistry to molecular biology, physiology, pathology, and circulation models. Particular examples include the effects of pathogens on water use and CO₂ uptake in leaves and roots,” says Farquhar.

“Gas exchange machines link different studies and scales from biochemistry to molecular biology, physiology, pathology, and circulation models.”

Graham Farquhar

 

In that sense, gas exchange instruments are essential to achieve translational outcomes. Susanne von Caemmerer, remarks that these instruments allow us to verify that we have achieved improvements in photosynthesis at the leaf level and then if this has any effect at the crop level. “This gives us an insight into leaf metabolism, linking photosynthesis to its underlying biochemistry,” she says. For example, if plants are grown under similar conditions, gas measurement allow scientists to study the range of photosynthetic capacity within a population of genotypes. “We then can use this knowledge to construct genetic maps of the trait, which are a powerful breeding tool for making crop improvements,” says Barbara George-Jaeggli, a Centre researcher.  

From big things, little things grow

Despite their importance for plant research, gas exchange instruments are relatively new inventions. Many plant scientists have been able to witness their origin and evolution. They have evolved from large laboratory building instrumentation to more complex, portable and precise instruments, like the ones scientists use today.

In the 1980s, John Evans used the gas exchange system in the Research School of Biology at ANU, built by Dr Chin Wong in 1970. “It had three different types of Infra-Red Gas Analysers (IRGA) and we used to record data with chart recorders and calculated all the parameters on the one main frame computer. We spent a lot of time calibrating these instruments, but as a result, we gained a deep understanding of the process. I even constructed a gas exchange system from instruments that were in the lab and used a portable IRGA to screen wheat genotypes in the field,” he says.

This same machine continues to be used by scientists at The Australian National University, but it is now difficult to guess its real age, as it has had several instruments attached to it during its lifetime. Although the new generations of gas exchange machines have several advantages such as being portable, lighter and produce faster results, they haven’t replaced the old ones as they have different purposes.

As Florian Busch, a Centre Researcher points out “with commercial machines everything comes in one box, you get results faster and measurements are easier. However, a home built machine, such as the “System 1”, is often more useful, as they are built for a specific purpose so you can get better and more appropriate information to address the questions you’re asking,” he says. “I have re-plumbed some of the commercial machines and hooked them up with other kinds of equipment, such as a mass spectrometer, so that I could get a greater set of data. I also built a special type of gas exchange machine, one in which I was able to control the mixtures of gases that I needed to test,” he says.

Getting faster results is a huge advantage of the modern versions. “When the LI6400 was first shown to us at ANU around 1990, I was skeptical about the benefits of integrating the leaf chamber together with the IRGA. It was only once I started using it that I appreciated how much faster one could make measurements,” says Evans.

Susanne von Caemmerer points out that the new generation of steady state infrared gas analysers have allowed the miniaturization of the instrumentation, together with new electronic transistor and computer technology. “The large systems had a versatility in the way leaf chambers could be used. The chambers often had better leaf temperature control and it was easier to control incoming air humidity. Now measurements can be made at much greater speed and an inbuilt computer will calculate the results immediately. This has greatly expanded the type of experiments that can be done,” she says.

To the future and beyond

As with any other technology, gas exchange machines are still evolving and there is room for improvement. Currently they are used all around the world, enabling a huge quantity of scientific innovation and discovery in the field of photosynthesis research (see box below). A clear example is the combination of gas exchange and carbon isotope discrimination measurements, which has led to a deeper understanding of the mesophyll conductance and its relation to temperature.

This technology continues being one of the best ways to access and unravel the complex photosynthetic process and our Centre researchers are recognised worldwide for their contributions in this area.

 

Gas-exchange instruments, in combination with other technologies are currently contributing to:

• •The ability to detect which wheat genotype has a larger photosynthetic capacity and if this happens in the enzyme Rubisco (Calvin cycle) or in the electron transport chain in chloroplasts.

•The establishment of a number of significant photosynthetic responses to growth under elevated atmospheric CO₂: C3 plants acclimate by reducing the content of the enzyme Rubisco while still maintaining  photosynthesis while C4 plants show little change.

•The establishment of the close linkage between photosynthesis and water use.

•The understanding of CO₂ sources and sinks in the biosphere.

•The understanding of how CO₂ diffuses through and is recycled inside the leaf.

•Understanding the effect of the environment on stomatal physiology.

•The development of photosynthesis models and testing of these models.