THE DEVELOPMENT AND APPLICATION OF BENEFICIAL BACTERIA AND FUNGI TO IMPROVE THE NUTRITION AND PHYTOSANITARY STATUS OF PLANTS AND SOILS IS AN INCREASINGLY POPULAR PRACTICE.
Examples include the Pseudomonas, Bacillus, Rhizobium, Trichoderma, Gliocladium and Clonostachys species.
The detection and quantification of these species following application allows us to monitor the product used and reliably establish its effects on the crop..
Study method 1
ENUMERATION OF VIABLE FUNGI AND BACTERIA ON SOILS, SUBSTRATES, LEAVES, STEMS AND FRUIT
It is impossible to detect or quantify the presence of bacteria or fungi applied to a plant on leaves, stems, fruit, roots or in the soil with the naked eye.
We have developed procedures to reveal and quantify the amount of micro-organisms that remain attached to plant or soil surfaces.
The results are expressed as colony-forming units (CFU)/gram or cm2 depending on the origin of the sample.
Study method 2
‘IN VITRO’ DETECTION OF PHYSIOLOGICAL ACTIVITIES
An increasing number of inoculants based on micro-organisms with different functions are being developed for application to soils. Examples include phosphorus (P) and potassium (K) solubilising bacteria. These are detected and quantified using samples of the soil or substrates where they have been applied. Dilutions are made in suitable media and after incubation the solubilisation index is calculated, which consists of measuring the solubilisation halo that has formed and comparing it with bibliographic reference data.
Phosphorus-solubilising bacteria. Transparent halos can be seen around the bacteria that have solubilised the phosphorus.
Potassium-solubilising bacteria. Note the yellow halo of colour change in the culture medium due to potassium solubilisation where the bacterial colony has developed.
Study method 3
STUDY OF ‘IN VITRO’ INTERACTIONS OF BENEFICIAL BACTERIA AND FUNGI WITH PATHOGENIC MICRO-ORGANISMS AND PRODUCTS WITH AGROFORESTRY APPLICATIONS (FUNGICIDES, PLANT EXTRACTS, INSECTICIDES, HERBICIDES).
These techniques consist of pitting two micro-organisms against each other in petri dishes (usually one is beneficial and the other is pathogenic) to study the effect of one on the other during their growth in the appropriate culture medium.
Different concentrations of a formulation in the culture medium containing the fungus or bacterium to be tested can also be used to determine the lethal dose, i.e. the concentration at which the micro-organism is killed by inhibiting its growth on the culture plate.
Many variations can be performed using the dual culture method as the basis, with the choice depending on the kind of product or micro-organism being tested.
Study method 4
ASSAYS TO EVALUATE THE EFFECTS ON GREENHOUSE PLANTS
This consists of inoculating beneficial micro-organisms on plants grown in pots under greenhouse conditions.
The pathogen is then introduced and the crop is maintained until the effects on the plants as a result of the interaction between beneficial and pathogenic organisms can be determined.
Example: “Study of the effect of mycorrhizal fungi on the control of Phytophthora in pepper plants”.
Pepper plants in pots inoculated with mycorrhizal fungi. The plants are infected with Phytophthora, a pathogenic fungus. Different treatments are carried out with the appropriate control samples:
- Treatment 1- Control plants without treatment
- Treatment 2- Plants inoculated with mycorrhizae
- Treatment 3- Phytophthora-infected plants
- Treatment 4- Plants inoculated with mycorrhizae + Phytophthora infection
At the end of the assay, the resulting information allows determination of effects between the two micro-organisms such as:
Weekly observations of plant condition, disease symptoms, degree of pathogen incidence, mycorrhization rate and amount of pathogen in soil (colony-forming units = CFU/g soil).
The following table shows the results obtained:
4. Mic + Phy
CFU Phytophthora per gram soil
2 x 106
2 x 102
COMMENTS ON THE RESULTS
We can infer that mycorrhization remained at a high level even when the plants were infected with Phytophthora. However, the pathogenic fungus reduced its concentration in the soil, at the same time drastically lowering the incidence of the disease, probably due to the presence of mycorrhizae in the roots which are able to protect the plants as well as nourishing them.
It may be concluded that the mycorrhizal fungus produces beneficial effects on pepper plant crops, allowing them to survive the attack of a pathogenic fungus.
This type of test, which can also be carried out with bacteria, plant extracts, etc., allows prediction of the effect of agricultural products under semi-field conditions and improved knowledge of dosages, application times, etc.
These experimental assays are very useful for companies manufacturing or marketing these formulations, with the data generally serving as a sales argument.