Knowledge grows on chickpea nematode nemesis

Flavonoids the focus for scientists as a defence against costly plant parasites

Weeds, Pests, Diseases
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Research paves the way for development of chickpeas tolerant to soil-borne pathogens.

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Australian National University Professor of Plant Science, Ulrike Mathesius, is researching flavonoids and their role in protecting chickpea roots against pathogen attack. PHOTO Nicole Dron

Australian National University Professor of Plant Science, Ulrike Mathesius, is researching flavonoids and their role in protecting chickpea roots against pathogen attack. PHOTO Nicole Dron

Compounds released from the roots of chickpeas are being explored in a bid to arm crops with in-built tolerance against the yield-robbing attacks of plant-parasitic nematodes.

Ulrike Mathesius, Professor of Plant Science at the Australian National University, is leading a team of researchers looking for new ways to tackle nematodes.

The microscopic roundworms under investigation include root-knot nematodes (Meloidogyne artiella, M. incognita and M. javanica), which together with cyst nematodes (Heterodera ciceri) and root lesion nematodes (Pratylenchus thornei) are estimated to reduce global chickpea yields by 14 per cent.

Plants under attack

A cross section through a healthy root with an ordered arrangement of vascular tissue in the centre of the root. Fluorescent flavonoids (blue coloured cells) accumulate in the root and are released into the soil. PHOTO Ulrike Mathesius

A cross section through a healthy root with an ordered arrangement of vascular tissue in the centre of the root. Fluorescent flavonoids (blue coloured cells) accumulate in the root and are released into the soil. PHOTO Ulrike Mathesius

Nematodes decrease chickpea vigour by feeding on the roots, reducing root nodulation and lowering the nitrogen-fixing capacity of the host plant.

"Nematodes cripple the root system, leading to water stress and nutrient deficiency, which reduces plant growth and yield," Professor Mathesius says.

"Secondary infections from fungi, such as Fusarium wilt and dry root rot, also occur when they infect the wounds created by feeding nematodes."

A cross section through a root infected with root-knot nematodes. The vascular tissue is disordered and the nematodes have caused the formation of giant cells from which the female nematodes feed. The different colours originate from different chemical structures of flavonoids formed in the diseased roots. PHOTO Ulrike Mathesius

A cross section through a root infected with root-knot nematodes. The vascular tissue is disordered and the nematodes have caused the formation of giant cells from which the female nematodes feed. The different colours originate from different chemical structures of flavonoids formed in the diseased roots. PHOTO Ulrike Mathesius

While Professor Mathesius has a background in root nodulation, the scope of her work has expanded since the Australian Research Council Hub for Legumes for Sustainable Agriculture was set up with support from GRDC in 2015.

She and her team are now focusing on exploring the relationships between nitrogen-fixing bacteria and parasitic nematodes on chickpea roots and how certain compounds these legumes release from their roots repel nematode attack. These compounds are known as flavonoids.

Flavonoids are signalling compounds released from the roots of all plants. In chickpeas, they influence a range of microbes in the soil by attracting them to their roots or repelling them.

"All plant species exude different flavonoid mixtures that have varied chemical structures," Professor Mathesius says.

"In legumes, one of their roles is to attract and activate rhizobia, the nitrogen-fixing bacteria that are essential for nodulation and nitrogen fixation.

"Without flavonoids, legumes would not be able to attract the right species of rhizobia to their roots."

Favourable flavonoids

A healthy root, left, and a root infected with nematodes, right. PHOTO Ulrike Mathesius

A healthy root, left, and a root infected with nematodes, right. PHOTO Ulrike Mathesius

While flavonoids are crucial for attracting rhizobia, the research has shown some of these compounds also help protect legumes' roots against plant-pathogen attack.

To explore which flavonoids are released from chickpea roots, the researchers extracted the compounds from the soil surrounding the roots (the rhizosphere) and used mass spectrometry to identify and quantify them.

The root-knot nematodes were then added to water and videoed to measure how fast they moved, by adding different flavonoids identified from the rhizosphere soil at varying concentrations.

Several flavonoids were found to increase or reduce nematode movement in a concentration-dependent manner.

Next, chemotaxis (repellent activity) assays were performed by drawing concentric circles on petri dishes and randomly dispersing the nematodes across the plates.

In the centre of each plate, a drop of the different flavonoids at varying concentrations was added and the resulting pattern of nematode movement away from the centre showed that some compounds (medicarpin, coumesterol and daidzein) could repel nematodes to varying degrees.

Interestingly, Professor Mathesius says, some of the flavonoids that repelled the nematodes were also those that attracted nitrogen-fixing bacteria. Others are known to be defence compounds against a number of different root pathogens.

Building resilient plants

In a proof-of-concept experiment using another legume species, the researchers developed transgenic plants that over-expressed the favourable flavonoids in their roots.

The plants were subsequently found to be more tolerant of nematodes and, pleasingly, produced more nodules for nitrogen fixation.

"We don't exactly know why yet and we haven't repeated this experiment with chickpeas," Professor Mathesius says.

"We are now screening 20 different commercial chickpea cultivars and extracting the flavonoids to see if there are correlations between the quantity of compounds produced and the tolerance or susceptibility of each cultivar to root-knot nematodes."

So far, she says, big differences have been discovered between the amount and type of flavonoids produced by the 20 cultivars. The work will be repeated for root lesion nematodes and cyst nematodes.

"Going forward, we hope to find a useful molecular marker to help chickpea breeders select genotypes that make high amounts of these useful flavonoids that repel nematodes and attract rhizobia," she says.

"Our next step is to repeat this glasshouse experiment in the field to explore how abiotic stress conditions change the flavonoid profile released, and whether those compounds that repel nematodes actually increase nodulation and nitrogen fixation under field conditions."

Going forward, we hope to find a useful molecular marker to help chickpea breeders select genotypes that make high amounts of these useful flavonoids that repel nematodes and attract rhizobia. - Australian National University Professor of Plant Science Ulrike Mathesius

Fungi focus

Professor Mathesius says preliminary work has also explored the effects of flavonoids on pathogenic fungi such as rhizoctonia.

"Another goal is to determine if the chickpeas that are more tolerant of nematodes are also more tolerant of soil-borne fungi," she says.

"Some of the flavonoids that inhibit nematode infection are known inhibitors of fungal attack, so we plan to test this in the future."

GRDC Research Code: US00083

More information: Professor Ulrike Mathesius, 02 6125 2840, ulrike.mathesius@anu.edu.au

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