Plants are always in contact with a huge number of microbes, above and below the soil surface. Some of these microbes are beneficial, others are harmful, while the vast majority are not known to have either positive or negative effects on plants. My work in Kamchatka focused on two plantmicrobe interactions that are beneficial for the plant: biological nitrogen fixation and endophytic fungi with anti-herbivore activity in grasses.

I. Biological N₂ fixation in different ecosystems

Nitrogen (N) is the plant nutrient needed in the largest amounts and N is often a limiting factor for plant growth and biomass production. This is because plants can use only a very few N compounds: ammonium (NH₄⁺), nitrate (NOH₃^–) and to some extent amino acids. These plant-available N sources occur only in very low concentrations in soil. However there is another possibility for some plants to obtain a useful source of N, and that is via symbiotic N₂ fixation.

Biological N₂fixation means that N₂in air is reduced to ammonia with the aid of a specific enzyme, nitrogenase. Only a small number of bacteria can form nitrogenase and thus fix N₂. A few genera of N₂-fixing bacteria can infect the roots of certain plants. The plant responds by forming a special structure, root nodules, where the bacteria live at a high population density and where they perform the N₂fixation process. Most of the nearly 20,000 species of legumes can form root nodule symbioses with bacteria collectively called rhizobia. Similarly, bacteria of the genus Frankia initiate root nodule symbioses with so called actinorhizal plants, which comprise some 200 woody species distributed among eight plant families. Well-known examples in northern areas are the genera Alnus, Myrica and Hippophaë. For simplicity, plants having symbiosis with N₂-fixing root nodule bacteria are called N₂-fixing plants.

In the root nodules the ammonia produced by the bacteria is immediately used by the host plant. Neighbouring plants and plants occurring at later stages in a plant succession also benefit from N₂fixation. Nitrogen originating from the fixation is delivered to the soil when dead leaves, shoots and roots are broken down in the soil. Another route is via urine and faeces when N₂-fixing plants are eaten by herbivores. Thus biological N₂fixation is a biological N fertilization.

Aim of the work in Kamchatka

N₂ fixation has been studied mainly in species that are used in agriculture. In Scandinavia the perennial clovers (Trifolium  spp.) and lucerne (Medicago sativa), and annuals such as pea (Pisum sativum), faba bean (Vicia faba) and vetch (Vicia sativum) are the most commonly used. Clovers and lucerne are characterized by a very high reliance on N₂ fixation; they often obtain more than three quarters of their N from N₂ fixation. It is not known if this is a characteristic resulting from a long period of domestication or if it is an inherent characteristic of these legume species. However surprisingly little is known about N₂ fixation in wild legumes in northern areas – do they have a similar high reliance on N₂ fixation for their needs? The question is of interest for our basic knowledge but it is also of interest to learn about wild legumes that may, in the future, become domesticated or incorporated into various agricultural systems. Learning about N₂-fixing plants in Kamchatka gives a unique opportunity to gain information about legumes that are not present in Sweden but are growing under more or less similar climatic conditions. The aim of my work in Kamchatka was therefore to collect as much information as possible about N₂-fixing plants growing in a variety of ecosystems. Data from the genera and species collected in Kamchatka will later be compared with corresponding data from Scandinavia.

Sites

My fieldwork was done at the four main sites described in Plant–herbivore systems in Kamchatka and during brief visits to the coast of the Sea of Okhotsk, southwest of Ust’-Bolsheretsk, and to the west coast of Karaginsky Island.

Field observations and collected material

Root nodules were present in all examined plants of all legume genera, namely Hedysarum, Lathyrus, Oxytropis, Thermopsis and Trifolium. The red-pigmented interior of the nodules indicates N₂-fixing ability. Alnus fruticosa, A. hirsuta and Myrica tomentosum also had root nodules.

Lathyrus japonicus was colonizing the outermost part of the sandy shore on Karaginsky Island. Thermopsis fabacea, with a pronounced southwestern distribution in Kamchatka, seemed to be a strong colonizer on bare ground and an invader into coastal heaths at Sea of Okhotsk. The species both had runners and a rich seed production. Members of the genera Hedysarum and Oxytropis were studied at the sites Ichinskaya Sopka Volcano, Karaginsky Island and Vilyuchinskaya Sopka Volcano. Both genera were represented on meadow terraces close to small creeks and rivers, but Oxytropis was also common on heaths at higher elevation. The clovers Trifolium pratense and T. repens were cultivated or seemed to have escaped from cultivation. They were observed in agricultural fields, on road-sides, along paths and in villages, but were not observed in areas without agriculture or settlements.

There are several methods to quantify N₂ fixation, but in field studies in remote areas and when the fieldwork is restricted to only single short stays at each site, the only possibility is to use the so called ¹⁵N natural abundance method. To apply this method I have collected samples of N₂-fixing species of Hedysarum, Lathyrus, Oxytropis, Thermopsis, Alnus and Myrica, as well as samples from a number of non-N₂-fixing herbs to serve as reference species. Chemical analyses will comprise N and other nutrients, while mass spectrometry is needed to determine ¹⁵N/¹⁴N isotope ratio. An estimate of N₂-fixation, expressed as the proportion of plant N derived from air, can then be calculated from isotope ratios in N₂-fixing species and reference species.

Different legume genera are infected by different rhizobia. This host specificity will be studied in the laboratory in those species from which I could collect seeds. If a “new” legume is to be cultivated, in an agricultural system or in soil restoration, it is essential to know which rhizobia are needed for a successful root nodule formation.

II. Endophytic fungi with anti-herbivore activity in grasses.

(In collaboration with Dr. Dawn Bazely, York University, Toronto, Canada) Fungi of the genus Neotyphodium live internally, as endophytes, in the aboveground plant parts of many grasses. The fungi produce alkaloids that are toxic to herbivores – insects as well as mammals. This toxicity causes illness and even death in grazing cattle and leads to great economic losses in USA and New Zealand. The presence of Neotyphodium is well-known in species of Festuca, Lolium and some other grasses in warmer areas. In contrast, there is only limited information about the presence of Neotyphodium in Sweden and other northern areas. The endophyte can be revealed by tedious microscopy or by immunological methods. Fortunately, it has recently been demonstrated that immunological methods can also be applied to dried grass samples. This opened a unique opportunity to study the presence of endophytes in grasses in Kamchatka. I have collected and dried samples from sites with cattle grazing and horse grazing, as well as sites where no recent grazing could be observed. Results from Kamchatka and from planned collections from Scandinavia and, preferably, northern North America, will extend our understanding of this type of plant–microbe interactions considerably.