By: Sean Westerveld, OMAFRA and Amy Fang Shi, OGGA

At this time of year, many unusual symptoms can show up in specialty crop fields. If a disease is suspected, samples can be sent to diagnostic labs for determining the cause. However, diagnostic reports are not always designed for growers to interpret, which can often lead to misunderstanding as to the potential causes of the symptoms. It is important to understand the limitations of diagnostic tests so unnecessary control measures are avoided.

Pathogen Identification

A proper identification of a pathogen causing a disease often involves four main steps:

Characterize the Symptoms – Comparing the symptoms to previous reports from the crop or similar crops (e.g., disease compendiums)

Examine Under the Microscope – Examining plant and pathogen parts under the microscope to look for characteristic fruiting bodies, spores and other structures

Plate on an Agar Media – Placing surface-sterilized sections of the plant on agar media to determine if a fungus or bacteria grows from the tissue. Since diseased portions of a plant can already be invaded by many secondary organisms, an apparently healthy section of the plant immediately adjacent to the diseased area is usually selected for plating. Once something grows on the agar, the pattern of growth along with microscopic examination of the structures are used to identify the organism.

Test Genetically – Modern diagnostics involves testing the plant materials, soil or fungus using genetic testing, such as PCR, to compare the DNA or RNA in the sample to previous studies and existing databases. Some organisms are not in the existing database of tests, and labs may not run tests that can identify every organism. This means that there may be pathogens present that the lab did not detect because they didn’t test for it. The test may be able to identify the exact species or even strain of organism or may only be able to identity a broader group of similar organisms. Please bear in mind, a genetic test can identify both living and dead organisms. So, an organism present in the report doesn’t always mean it is alive and the cause of the disease.

Simply doing these steps is not enough. It is also important for a diagnostic lab to compare the results to previously reported diseases of the crop, gather details of the pattern of disease in the field and to ensure the identified organisms make sense for causing the symptoms reported. Simply finding that an organism is present does not mean that it is causing disease. There are numerous species of fungi, bacteria and other microorganisms in the soil and in the plant that either do not cause disease or actually benefit the plant in some way. In plant pathology, another step in a diagnosis is conducting a series of inoculations and isolations called Koch’s Postulates, but this would be too expensive or time-consuming for a diagnostic lab to undertake.

Diagnostic Reports

What should be reported to growers is the best guess of the lab as to the cause of the symptoms based on all the steps of a diagnostic test, but sometimes the lab just reports what was found and does not suggest a cause of the issue. It is important to realize that many issues in the field are abiotic, meaning they are caused by environmental (e.g., wind damage, heat stress, frost injury, pollution), nutritional (e.g., nutrient deficiencies and toxicities), pesticide application (e.g., herbicide drift, pesticide burn) or mechanical injury (e.g., damage from equipment or workers). Insect or vertebrate pests can also cause disease-like symptoms. When there is damage to the plant, weak pathogens can invade the tissues and will often show up on a diagnostic report.  It is important to recognize that organisms isolated from damaged tissues may not be the primary cause of the problem. To better understand when an organism isolated from a disease sample is likely the primary cause it is necessary to understand the different feeding types of microorganisms in the field:

Definitions

Pathogen (Parasite) – an organism that feeds in or on another living organism from which it extracts nutrients.

Secondary Organism (Saprophyte) – an organism that feeds on dead and decaying matter from which it extracts nutrients. They are considered “secondary” because they only come into diseases samples after a primary pathogen has killed the tissues.

Within these two types of feeding habits are some sub-categories (refer to Figure 1):

Figure 1. Diagram of the different types of feeding habits of organisms associated with diseased plant materials.

Definite Pathogens (called obligate parasites in plant pathology) – a type of pathogen that can only live on another host, and therefore, if present in diseased tissues is most likely the cause or a cause of the damage. Examples of diseases caused by definite pathogens include powdery and downy mildews, rusts, smuts, white rusts, all viruses, and clubroot (Figure 2). When an obligate parasite is found in plant tissues, it is most likely causing disease symptoms, and the symptoms should be relatively easy to compare to previous reports and compendiums. When these are present, growers can be confident that the diagnosis is accurate, although other factors may still be involved in making the crop more susceptible to the disease. Note however, that some viruses may be present but not causing disease symptoms. Since definite pathogens are often the cause of major and aggressive diseases, there is also value in ruling these out in a diagnostic test.

Figure 2. Pseudoperonospora cannabina, the cause of hemp downy mildew, is an example of an organism that is a definite pathogen.

Occasional Pathogens (called facultative parasites in plant pathology)– organisms that are usually secondary but that can become a pathogen under certain circumstances (e.g., favourable weather, weakened plant tissues due to another cause) (Figure 3).  Many of these kill parts of the plants and then feed on the dead material. When an occasional pathogen is found in the diseased sample, its presence alone is not sufficient for a proper diagnosis. The lab should confirm that it is causing disease by isolating it from diseased samples (i.e., not entirely relying on genetic tests) and ensuring that the symptoms match previous reports. For diseases of above-ground plant tissues, this is often relatively easy since symptoms are usually unique and specific to the crop. However, this becomes more difficult for root diseases and those specialty crops that do not have many previously published reports. In this case, the lab will make a best guess based on symptoms in other similar crops.

Figure 3. The coral-coloured fruiting bodies on this hazelnut twig are caused by Nectria. Although Nectria can be pathogenic to woody plants, it is usually secondary and only appears after the plant is damaged by something else (e.g. another disease or winter injury).

Occasional Secondary Organisms (called facultative saprophytes/saprobes in plant pathology) – a type of organism that normally lives as a pathogen but can survive on dead or decaying matter for at least part of its life cycle (Figure 4). Although it is possible for occasionally secondary organisms like Phytophthora to be living on decayed matter, they are also likely to be causing disease if present in a diagnostic report. However, it is possible that a genetic test of the soil will pick these up even if they are not causing disease. The lab will usually confirm that the symptoms on the plant match previous reports on that pathogen before making the diagnosis and/or isolating that organism from plant tissues plated on agar media.

Figure 4. Phytophthora nicotianae, the cause of Phytophthora root rot on lavender, is an example of an occasional secondary organism. If it is found in a soil or root sample, it is most likely causing disease.

Definite Secondary Organisms (call obligate saprophytes in plant pathology)– a type of secondary organism that only lives on dead and dying tissues, so cannot cause disease. They may be present in the dead portions of a diseased sample or as common contaminants. These organisms should not be reported in a diagnostic test but may show up in a PCR multi-test or as a contaminant in plated samples.

Interpreting Diagnostic Reports

Most of the issues with misinterpretation of laboratory reports comes with occasional pathogens. The real difficulty is when they are found in diseased roots and other below-ground plant parts. Some fungi such as Fusarium and Pythium are widespread in virtually all soils and will routinely invade dead and dying parts of plants. For example, we have received diagnostic reports on hundreds of soil and root samples over the years for ginseng research and in various other specialty crops and at least one species of both Fusarium and Pythium have been identified in 100% of those samples. In some crops and some circumstances, they may cause disease, sometimes severe, but it is impossible to determine if they are the primary cause based on their presence alone. Symptoms can sometimes help but are often less distinct in below-ground parts, and most specialty crops will not have previous reports for a comparison of symptoms. It is very important when dealing with facultative parasites that the sample be submitted to the lab in a timely manner and include both healthy and damaged tissues. To be more confident that they are the cause of the symptoms requires a combination of:

1. Ensuring the sample submitted to the lab has both living and damaged tissues and that they arrive at the lab in a timely manner. Heavily degraded or rotted samples are less likely to reveal the real culprit because secondary/saprophytic organisms will take over the sample. The lab should attempt to isolate the pathogen from healthy tissues adjacent to the damage when plating out the sample.
2. Determining if the species found is common or uncommon in the soil. For example, a Fusarium species not normally found in root samples is more likely to be causing disease than a species found in every root and soil sample. The lab technician can base that on their own experience, but a grower would need to submit numerous samples over time to know what is common or not.
3. Determining if the symptoms make sense for that pathogen. For example, Fusarium wilts are caused by Fusarium invading the vascular system and blocking the flow of water to the top of the plant. This can lead to distinctive discolouration of the vascular system. If that symptom is found in conjunction with the presence of a Fusarium species, then there can be more confidence in the diagnosis. The pattern of damage in the field can also be characteristic of certain pathogens. For example, patches of damage are more likely to be caused by a root disease like Rhizoctonia than foliar pathogens or abiotic issues like spray burn or nutrient deficiencies.
4. Determining genetically if a strain or special form of the pathogen species is present that is known to be specific to that host crop. This is not always possible without extensive genetic studies.
5. For specialty crops in particular, ensuring the lab compares the results to the crop or similar crops. It is best to ask the lab before submitting the sample if they have the appropriate tests for your specialty crop.

Other Potential Causes of Disease-Like Symptoms

Without some other way of confirming when a facultative parasite is causing a symptom, the presence of these organisms in a diagnostic report does not help in determining the cause, especially for root diseases. It is best to look at other potential causes before determining how to manage the issue. The following are the most common causes of symptoms that can be mistaken for diseases in a diagnostic report:

1. Winter injury. For perennial crops, winter injury can weaken the plant or damage the vascular system. The effects of this damage are often delayed and may not show up until mid-summer. These symptoms can be mistaken for root or stem diseases.
2. Spray damage. Damage from a pesticide application, drift from a neighbouring field, or herbicide residues in the soil are common. Many herbicides are systemic and may weaken the entire plant. Symptoms can range from yellowing, death of sections of the plant, stunting, twisting or symptoms similar to viruses or nutritional disorders. Spray history and the pattern of damage in the field is the best clue when a pesticide damage has occurred.
3. Nutritional issues. Nutrient deficiencies and toxicities can kill portions of the plant or cause chlorotic patterns in the leaves. Comparing foliar tissue samples between a healthy and a damaged part of the field can help to determine if a nutritional issue is the cause. Nutrient issues should be relatively widespread in the field or at least confined to a distinct area of the field (e.g., low or high area).
4. Moisture stress. Moisture stress (either too much or too little) can result in wilt, root diseases, nutritional issues or dieback.
5. Soil insects and vertebrate pests. Anything that damages the roots or bark can lead to disease-like symptoms and the lab may not be able to diagnose the issue if there is no obvious chewing damage and the pest was not collected with the sample.

It may be helpful to explain more details to the lab technicians about pattern and extent of damage in the field.

If you receive a diagnostic report and are unsure how to interpret it, first contact the lab to determine if they can provide more context. Because you are paying for a diagnosis, they are often motivated to provide a cause of the symptom, but they should be able to provide their confidence level that what they found is the cause of the issue. You can also reach out to an OMAFRA specialist who may have more experience with diagnostic reports and symptoms for a specific crop.