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These days a lot of studies are investigating mycorrhizas, which are associations between plants and certain soil fungi called arbuscular mycorrhizal fungi (AMF). Mycorrhizas have the potential to improve plant productivity in a range of ways, from increasing a plant’s uptake of nutrients, to improving its tolerance of stresses like soil salinity and disease. 

With those exciting possibilities, it is no surprise many mycorrhizal studies target annual crops like cereals and soybeans. Now Jonathan Bennett, an assistant professor at the University of Saskatchewan (USask), is delving into forage mycorrhizas.

Bennett and his research group are working on a series of studies to see if they can enhance forage plant responses to AMF and adjust AMF communities to boost benefits to forage crops.

He is collaborating on these studies with several other researchers, including Bill Biligetu at USask, Monika Gorzelak with Agriculture and Agri-Food Canada (AAFC) at Lethbridge, Sean Asselin with AAFC at Swift Current and J.C. Cahill at the University of Alberta.

A bit about mycorrhizas
In a mycorrhizal relationship, the fungus colonizes a host plant’s root, growing inside the root and developing thread-like fungal arms known as hyphae. The hyphae grow beyond the root into the soil to acquire nutrients for the plant. In exchange, the plant gives the fungus carbon sources like sugars and fats the fungus cannot produce on its own. In other words, AMF must have a plant host – they obligate plant symbionts.

Communities of multiple AMF species occur naturally in most soils, including agricultural soils, and AMF can form relationships with most types of land plants, including most crop species. Natural AMF communities are adapted to their local growing conditions – soil, climate, plant species and so on – so the fungal species and strains in AMF communities vary from place to place. 

Bennett notes some agricultural practices, such as intensive tillage and heavy fertilizer use, can degrade the natural AMF community, changing which fungal species are present and/or their abundance. So, the AMF community in a crop field may not be as beneficial to crops as it could be. 

Improving mycorrhizal interactions might be especially helpful for sustainably increasing forage production. For instance, mycorrhizas are important for providing phosphorus to plants, and Bennett notes that forage legumes are often phosphorus-limited. As well, perennial forages are often grown on marginally productive lands that may be more prone to problems like drought or soil salinity, and mycorrhizas can help plants deal with those problems.

Improved forage varieties
A recent study led by Bennett is aimed to determine if alfalfa and sainfoin plants can be selected for better mycorrhizal interactions. This is a first step toward the possibility of breeding alfalfa and sainfoin varieties that are especially good at forming mycorrhizas for increased forage productivity.

Bennett’s group obtained four-year-old alfalfa and sainfoin plants from breeding plots operated by forage breeder Biligetu. They transplanted those plants – including the soil around the roots – into pots and grew the plants in the greenhouse.

They evaluated the roots of the plants to assess the amount of colonization by the AMF naturally resident in the soil. Then they divided the plants into two groups: high-colonization plants and low-colonization plants.

They put all the high-colonization plants in one cage and the low-colonization plants in another cage. Next, they put some bees in each cage for cross-pollination among the high-colonization plants and among the low-colonization plants.

Using the resulting seeds from the high- and low-colonization lineages, Bennett’s group grew the seedlings in the greenhouse. Then they transplanted those seedlings for a field trial near Saskatoon to see if colonization by the resident AMF community would differ between the two lineages.

“It turned out that the offspring of the high-colonization alfalfa lines do tend to be more heavily colonized than the offspring of the low-colonization alfalfa lines, and the same with sainfoin,” says Bennett.

Furthermore, heavy AMF colonization appears to be a good thing for forage yields. He explains that mycorrhizal fungi form structures inside the root for active nutrient exchange: arbuscules, which are where the plant and the fungus exchange nutrients; and vesicles, which are the storage organs for the fungus. “Our data suggest the plants with more active mycorrhizal associations and more of these nutrient exchange and storage structures tended to be bigger when we grew them out in the field.”

Better AMF communities
Bennett and his group are currently conducting a study using mycorrhizal fungi cultured from AMF collected in saline areas in natural Prairie grasslands. They want to find out if these grassland AMF will improve salinity tolerance in three different forage hosts: a salt-tolerant alfalfa variety; tall wheatgrass, which is a highly salt-tolerant perennial grass; and a fairly salt-tolerant forage barley variety.

They are interested in AMF from natural grasslands in part because of the results from a previous trial where they applied a commercial AMF inoculant to various alfalfa varieties. “We didn’t see any real benefit to alfalfa growth from the commercial inoculant. In fact, it made the plants smaller in a lot of cases,” notes Bennett. 

Like many commercial inoculants, the one used in that earlier trial contained a single AMF species called Rhizophagus intraradices, rather than a community AMF species. Furthermore, like any mycorrhizal fungus, a commercial AMF strain is adapted to its own set of conditions, and its effect on crop performance will be influenced by the new growing conditions it encounters, including the crop type. 

“If you are trying to use some mycorrhizal fungi to improve plant growth, then you need to have the right mycorrhizal fungi for that environment,” Bennett explains. “Most of the cropping areas in Saskatchewan and Alberta used to be natural grasslands before they were plowed. So, natural grasslands may have mycorrhizal species that are well adapted to the local conditions, especially for perennial forages, which have more time [than annual crops] for those fungi to get established.”

Bennett’s group collected soil samples from 33 different grassland sites across Alberta and Saskatchewan. Those 33 sites included 16 paired saline and non-saline sites and one unpaired site. 

“My thought was that, in cropland, saline areas tend to be bare patches, with maybe kochia and foxtail barley and occasional crop plants. Salty areas in crop fields are probably fairly depleted in terms of the abundance of mycorrhizal fungi,” he says. “However, in natural grasslands, salty patches are pretty filled in with a diverse complement of plant species so they might be a good source of AMF communities that could confer salinity tolerance.”

The group brought the 33 samples back to the lab and cultured the AMF in each sample to prepare the fungal inoculum. Since mycorrhizal fungi must have a plant host, they grew the inocula using a trap culture. A trap culture involves a plant host species that is known to associate with most AMF species and to really promote their growth. As a result, the trap culture soil gets lots of AMF spores, hyphae and colonized roots, and can then be used as an inoculum in subsequent experiments.

Bennett’s group ran two greenhouse trials with the three forage species treated with the different grassland AMF inocula. The forage plants were grown in salt-treated soils that were moderately saline. 

“Some of the inocula did really well in improving salinity tolerance, particularly in barley where the responses were strongest. Certain inocula tripled the size of the barley plants compared to non-inoculated barley in the salt-treated soils,” he notes.

However, he cautions, “Not all the grassland inocula made the forages grow better in our greenhouse experiments. Roughly 20 per cent of the cultures we tried showed some sort of benefit.”

In the coming months, his group will be identifying the microbial species in the different inocula to see which species tend to be associated with the greatest improvements in forage plant performance. 

Looking ahead
One way these mycorrhizal studies might benefit forage production is by helping breeders to improve forage varieties. “Ultimately, we’d like to be able to breed forage varieties that work a lot better in harsher conditions with fewer inputs, whether that is with less phosphorus applied or if they’ll grow better in salty areas or under other stresses like drought,” says Bennett.

“Our research shows it is possible to select for lines that are highly mycorrhizal. Now the question is just how beneficial would it be to breed forage varieties for this trait? We’ll have to answer that question down the road.”

Another possible benefit is that their research findings could help inform forage producers who want to try culturing their own AMF inoculum from their own grasslands. To achieve that objective, his group will first need to figure out how to identify locations in natural grasslands that are likely to have really beneficial AMF communities. 

Bennett’s group has already made a start on this objective. When they collected the 33 grassland soil samples, they also collected data on the plant species composition, soil conditions, soil nutrients and other relevant characteristics at each sampling location. The researchers will be using various statistical methods to try to determine which particular growing conditions are associated with the best AMF communities for use with forage crops.

The practicalities of collecting, culturing and applying your own natural grassland AMF inoculum for field-scale forage production would also need to be worked out. For instance, how would you collect sufficient grassland soil for your inoculum production without degrading the grassland? Bennett says, “I would never want to encourage someone to be digging up a natural grassland, because we know these grasslands take a really long time to recover.” 

A further potential benefit is that Bennett and his group might identify an AMF inoculum with commercial potential for forage crop applications. If they are successful, Bennett thinks they would likely work with a commercial inoculant partner to see what would be the best strategy for making the inoculant available to Prairie forage growers.

There are still many questions to be answered to determine how beneficial the findings from Bennett’s forage mycorrhizal research are likely to be. He really appreciates that the Alberta Beef Producers, the Saskatchewan Cattlemen’s Association and the Natural Sciences and Engineering Research Council of Canada are willing to give this pioneering research a chance.