Researchers reduce trifluralin resistance

Researchers reduce trifluralin resistance

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 AHRI communication leader Peter Newman said the researchers strongly suspect that the absence of one or a number of P450s is what is causing prosulfocarb resistance.

AHRI communication leader Peter Newman said the researchers strongly suspect that the absence of one or a number of P450s is what is causing prosulfocarb resistance.

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Unexpected results changed the direction of the study.

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TRIFLURALIN resistance has disappeared in two populations of ryegrass that are being studied by researchers at the Australian Herbicide Resistance Initiative.

Researchers Roberto Busi and Danica Goggin started with trifluralin resistant ryegrass and then bred prosulfocarb resistance using recurrent selection over several generations.

As the ryegrass became more prosulfocarb resistant it became susceptible to trifluralin, which is known as negative cross-resistance and it's extremely rare.

Dr Busi said the study was one in which the results were unexpected and not the focus of what they were originally doing.

"My aim was to select for resistance to Sakura and Boxer Gold in ryegrass, but I found that when I sprayed ryegrass with prosulfocarb, which is the main active of Boxer Gold, the subsequent progeny of those plants lost trifluralin resistance," Dr Busi said.

"That was a surprise and the study since then has basically been rewinding and trying to understand why that happened."

Dr Busi used recurrent selection in the lab to 'breed' resistance to several herbicides, including pyroxasulfone (for example Sakura) and prosulfocarb (for example Arcade).

Recurrent selection involves spraying a population of ryegrass with a herbicide, then ensuring that the survivors cross-pollinate with each other and collecting the seed.

In each case at least 100 plants were sprayed and at least 10 survived which were isolated so they could cross only with each other.

For the study, the results of which have been published in a paper titled 'Loss of trifluralin metabolic resistance in Lolium rigidum plants exposed to prosulfocarb recurrent selection', all the populations that were used were coming from the field and were resistant to trifluralin.

"The experiment went on for two years, so I did a recurrent selection with prosulfocarb and at the end of the two year study, all of the material generated from the field and under controlled conditions was screened with either prosulfocarb or trifluralin," Dr Busi said.

"We monitored what happened to the initial field resistance to trifluralin and found that when we used prosulfocarb, the progeny of those ryegrass populations is less and less resistant to trifluralin.

"We knew that these populations were resistant to trifluralin and that capacity was lost after prosulfocarb use, so we went to the lab to quantify and measure the ability of those ryegrass seeds and seedinglings to detoxify trifluralin."

From that, Dr Busi and his team found that what they saw in pods at the whole plant level was also confirmed in the lab as those plants, after two years of recurrent use of prosulfocarb, were less able to metabolise trifluralin.

AHRI communication leader Peter Newman said Dr Coggin teamed up with Dr Busi to measure the metabolism of trifluralin as well as study the P450 enzyme activity using the P450 inhibitor, phorate.

"One of the main mechanisms of trifluralin resistance is metabolism based resistance by P450 enzymes, we know this because when researchers use a P450 inhibitor such as phorate, they can switch off the resistance and that is exactly what Roberto and Danica did in this study," Mr Newman said.

"For example, for the SELR68-P0 population, pre-treating the ryegrass seeds with phorate reduced trifluralin survival from 41 per cent to 4pc, turning off the P450s with phorate turned off the resistance."

Mr Newman explained that when prosulfocarb was first taken up by a ryegrass plant, it's not herbicidal.

"Prosulfocarb must be activated within the plant by a P450 enzyme to become herbicidal, this is what we call a pro herbicide," he said.

"P450 is short for P450 mono-oxygenase, they add one oxygen to a molecule and once inside the plant, a P450 adds an oxygen to prosulfocarb and we have an active herbicide."

Given prosulfocarb needs activation by a P450, if they disappear, prosulfocarb doesn't get activated and it doesn't work anymore.

In this study, Dr Coggin and Dr Busi treated prosulfocarb susceptible ryegrass with the P450 inhibitor phorate and stopped prosulfocarb from working.

"The researchers strongly suspect that the absence of one or a number of P450s is what is causing prosulfocarb resistance," Mr Newman said.

"If the same P450 that is responsible for trifluralin resistance disappears, if it's the same one that is causing trifluralin resistance, then the trifluralin resistance disappears."

While the discovery made by Dr Busi and Dr Coggin is groundbreaking, it's important to remember that there's always another resistance mechanism and this phenomenon is unlikely to happen with every population of ryegrass.

"The message is to continue to mix trifluralin with Arcade (prosulfocarb), Boxer Gold (prosulfocarb + metolachlor), Sakura, Avadex, etc.," Mr Newman said.

"If trifluralin resistance and prosulfocarb resistance are mutually exclusive in a number of ryegrass populations then the mix of these herbicides makes a lot of sense, from both a resistance and efficacy point of view."

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