Fungi get everywhere. Everywhere. It’s no surprise that they associate with plant roots in the soil, but they don’t just live in and around roots. They grow in plants above ground too. Some of these fungi are pathogens that attack the plants, but not all.
Some fungi provide defences for the plant but producing toxins that attack insect herbivores. If they don’t make these defences themselves, they can at least trigger the plant to produce those chemicals. There are plenty of fungal infections that can benefit a plant.
Between these good and bad fungi, there’s a third group of unspecialised fungi. These travel on the wind as spores from plant to plant, settle down a while and then move on. It has been thought that these fungi had no interest in plant-herbivore interactions. A new paper, Meta‐analysis of the role of entomopathogenic and unspecialized fungal endophytes as plant bodyguards, by Professor Alan Gange and colleagues, sets out to tackle that assumption.
It’s an important question because there is a lot of these fungi, and they’re widespread. Prof Alan Gange said: “It is exceptionally rare for any plant not to be colonised by at least one species of fungus. However, most plants don’t seem to harbour very many at any one time – it’s probably between about 5 and 10. Thus, even if the total number of fungi found across a population of plants might be high, they don’t all occur within one plant. An example is Cirsium arvense – we have found 64 different species living as endophytes within this plant, but the most we have ever found in one plant is 10 and the mean per plant is 5.6.”
One approach to studying the effect these endophytes have on insects is to set up an experiment. Quite a few people have done this. A meta-analysis builds on this by examining all these studies and seeing if any patterns emerge by pulling the studies together. This is fine as far as it goes, but the GIGO principle applies. How do you know you’re not getting a biased result by selecting a biased selection to start with.
To tackle this, the team had a three-step approach. First, they compared published results against unpublished results from their own labs. There’s a well-known bias in publishing that it’s easier to publish results with a significant result than inconclusive studies. The problem is that if the reality is that there’s not much effect, then you’d expect a lot of inconclusive results. As it happens, they found close agreement between their unpublished studies and what they saw in journals.
The next test was to see if there was bias in the results. Chance can interfere with results, but if there’s no bias, then results will be scattered around the ‘real world’ figures. If there is bias, then you should see people favouring results on one side rather than another. This test produced mixed results, with there appearing to be some bias – but this was due to some extent to the nature of the statistical tests used. So they added another test.
Gange and colleagues had significant results from their analysis of publications and their own unpublished results. But what if the team had missed other unpublished results? Could another three unpublished experiments, lying in a drawer somewhere, change what looked significant into the kind of thing you’d expect by chance? They conducted another statistical check to work out how many missing publications they’d need to negate their results. This final test showed you’d need a staggeringly large number of unaccounted studies to damage the conclusions.
So what did they find?
Unsurprisingly, they found entomopathogenic fungi harmed insects. What did surprise them was how. A simple experiment would be to add fungi to a plant and then see what happens. But when people examined leaves eaten by insects, they didn’t find any fungi. Nonetheless, the fungi were having a strongly negative effect on the insects. It seems fungi were capable of influencing the plant without having to be everywhere in the plant.
There was another surprise in the data. The nonentomopathogenic fungi also had a negative effect on insects – despite not being poisonous to insects. I didn’t expect to see that, nor did Prof Gange. “Yes, this surprised me – the fact that these fungi which are usually found in soil as saprophytes could exist asymptomatically within plants and have effects on the insect herbivores that eat those plants. We think the explanation is a chemical one – we know that when one of these endophytes infects a plant, significant chemical changes occur. Thus, while the fungus itself appears not to grow systemically through the plant, the chemical changes are systemic, and this is what affects the insect feeding.”
Which insects are the most affected by the fungi? Would it be the herbivores who specialise on a plant or more generalist browsers? It wasn’t immediately obvious before the search. After all, endophytes could also specialise in certain plants, and so help defend against a familiar pest. In fact, it was the insects that generalised that fared worst. The team concluded that a generalist is always landing on a new chemical cocktail produced by endophytes, so never develops countermeasures. It also pays to be a leaf-miner, to avoid the worst effects. For some reason, sucking the sap led to bigger negative effects from nonentomopathogenic fungi.
Reading so many studies also allowed the team to compare procedures. Prof Gange said: “The procedure that creates the biggest effect is infection of the seeds. This is counter-intuitive as the vast majority of these fungi infect leaves via air-borne spores. It is not surprising that many experiments have tried to infect leaves with spores and then measured effects on insects – and these experiments show very little effect. I think the reason is that the fungi only successfully invade leaf tissues if the air is very humid, allowing spore germination on the leaf surface. Thus, unless you recreate the conditions of a heavy dew, you won’t infect your plant. However, seed treatments mean the fungus is in intimate contact with the seed in a humid environment, so they go in easily. So the fungus enters the seed, and then the seedling, and so the plant is chemically altered from the start. It means that when the plant grows up, it has this chemical protection – the ‘bodyguard effect’.”
The ‘bodyguard’ employs fungi that you’re probably already eating whenever you have a salad. But would it be possible to give fungal protection to seeds systematically, to reduce pesticide use? Prof Gange replied: “I sincerely hope so – it is easy to encapsulate seeds, and I see no reason why we could not incorporate certain fungi into those seed coatings.”
Getting the right fungi could take some work, as endophytic fungi aren’t well studied. Prof Gange said: “They are quite poorly known, ecologically speaking. However, all the species we find as endophytes have alternative lifestyles – many are saprophytes in soil or are pathogens of other plants, so they are reasonably well known, taxonomically.”
So how would you carry out a PhD to work on endophytes and plant defence? Prof Gange said: “I think it should focus on the mechanisms because what I have said above is speculation. Questions like: 1) does the fungus acquire carbon (or other nutrients) from the plant? 2) are the chemicals produced fungal-induced or fungal produced? 3) why are sucking insects affected most – is it chemical transfer in the phloem? and 4) can we develop commercial seed treatments across a range of plant species? It would form a great thesis.”