How histology can help understand the soybean “anomaly” in the field

The term "anomaly" became widely used by rural producers in the BR-163 region, especially after the 2018/2019 harvest, to describe a new phytosanitary situation affecting soybeans in their reproductive stages. According to the Portuguese dictionary, "anomaly" is defined as an abnormal, irregular, or unusual condition.

Therefore, farmers and technicians began using this term to describe atypical symptoms observed from the R5.1 stage onward, such as pod rot and stem breakage, which did not fit into the traditionally recognized diseases of the crop. The use of the term is justified by the unusual manifestation of these symptoms, which deviate from the expected physiological pattern for soybeans.

It is estimated that, depending on the climatic conditions of the harvest and the cultivars used, the damage caused by the so-called soybean “anomaly” in the mid-north region of Mato Grosso could reduce productivity by between 16% and 30%, which represents a potential loss of up to 59 million bags.

Symptoms and stages

Initial symptoms usually appear between the R5.1 and R5.5 stages, when the seeds are still forming and can be felt. At this point, slight changes are observed in the pods (Figure 1A), which, despite their apparently normal external appearance, already show internal damage. As the process progresses, lesions that may darken on the inside of the pods (Figure 1B) are observed, often associated with the presence of fungal mycelial growth on the still-developing seeds.

As the condition progresses, the seeds begin to exhibit wrinkled, uneven seed coats, with dark coloration and apparent loss of specific mass (Figure 1C), although some of the internal mass may still remain preserved (Figure 1D). These changes directly impact the physiological and health quality of the seed, compromising the productive potential of soybeans.

The distribution of symptoms within the plant canopy and among the seeds does not follow a defined pattern. Symptomatic pods can appear in different parts of the plant, and affected seeds occur randomly within the pods—that is, not all seeds in a given pod will necessarily show symptoms (Figure 2). At the R6 stage, even with the pod still closed, signs of early seed germination can be observed, which can lead to pod opening at more advanced stages (Figure 3).

Laboratory analysis

Laboratory diagnostics performed on symptomatic tissues revealed the presence of different pathogens, such as Fusarium spp., Diaporthe spp. And Colletotrichum spp., suggesting a possible interaction between biotic and abiotic factors in the etiology of the disease. Although some studies attribute the cause of seed rot to species such as Diaporthe ueckeri e Diaporthe longicolla, this hypothesis is still not consensual within the scientific community. The debate remains open, and there are different interpretations regarding the origin and agents involved in the observed symptom complex.

Our research group, for example, isolated a fungus belonging to a genus whose species has not yet been described in the international scientific literature, and molecular data indicate that it is a distinct phytopathogen from those traditionally associated with the disease. These findings reinforce the need for further investigation and collective knowledge development on the topic, especially given the complexity of the symptomatic picture observed in the field.

In addition to symptoms related to seed rot, our field monitoring—conducted since sowing over the last four harvests—also recorded the occurrence of hypocotyledonary lesions and stem splitting (Figure 4). Although some professionals consider these manifestations to be normal physiological variations, this interpretation is not supported by the specialized botanical literature, which reinforces the need for further investigation into their origin and impact on plant development.

Therefore, it's crucial that these symptoms be treated with due technical rigor and not dismissed as mere morphological variations. Correct diagnosis is essential to guide effective management strategies and mitigate potential productivity losses associated with this type of occurrence.

In this context, researchers from the University of São Paulo (USP) and the Federal University of Mato Grosso (UFMT) joined forces to understand more deeply how the symptoms of the soybean "anomaly" affect different plant tissues. As an initial result of this collaboration, an expanded abstract was published at the 39th Soybean Research Meeting (Leonardi et al., 2024), presenting histological and ultrastructural approaches to analyzing the problem.

For this type of investigation, advanced microscopy technologies are required, which enable comparative visualization between healthy and diseased tissues through high-resolution images, with the use of specific dyes that interact with distinct chemical groups, allowing the highlighting of regions and compounds of interest (Marques; Soares, 2021).

Thus, samples of soybean plants (Glycinemax) were collected at Fazenda 3 Irmãos, located in Sinop (MT) and taken for analysis at the “Luiz de Queiróz” School of Agriculture, where they were prepared - fixed (Karnovsky, 1965), stained with Toluidine Blue dye and analyzed under a light microscope and scanning electron microscope.

Sample preparation

Initially, the plant samples underwent a ketonic dehydration series in different acetone concentrations (30%, 50%, 70%, 90%, and 100%) and were gradually immersed in resin (Historesin) to ensure complete infiltration of the material, allowing for good microscopic visualization. After this process, the samples were placed in silicone molds, where they polymerized as the resin hardened, forming blocks. These blocks were cut to a thickness of 7 μm using a Leica 5460 microtome to produce glass slides. Subsequently, the slides were stained with dyes and viewed under a Zeiss AxionVision light microscope (Figure 5).

Research results

Images taken using microscopy and plant histology techniques for the stem region allow for the identification, at the tissue level, of symptoms observed in the field through the formation of cracks in the plants (Figure 6).

It is possible to analyze that plants that did not develop cracking presented an organized outermost layer (epidermis), with a layer of intact cells and the presence of fibers external to the phloem region, without compromising the conducting vessels (Figure 6A). However, in plants with symptoms of "anomaly," cracking develops, resulting in the loss of the epidermis, and, in response to this damage, the formation of a healing meristem is observed, as an attempt at recovery by the plant (Figure 6B).

Furthermore, in the cortex, the cells showed hypertrophy (increase in size) and hyperplasia (they multiply more than normal). Finally, cell death, plasmolyzed cells, and the presence of fungal hyphae that developed in the spaces between the cells were noted.

The closing thought

Many questions remain regarding the "anomaly," as well as the pathogen-host interaction, and how it affects productivity. Furthermore, there are doubts related to the plant's response: whether it is exclusively due to pathogen attack, or if there is another stress causing the opening of cracks at the base of the plants that serve as an entry point for fungal infection.

Significant progress has been made in molecular and morphological analyses, with the aim of accurately identifying the pathogen(s) involved in the observed symptomatic picture.

Accurate etiological characterization is essential to support more effective management strategies and mitigate productivity losses recorded in the field. In this context, histology and histochemical techniques emerge as valuable complementary tools, allowing for a detailed analysis of pathogen-host interactions in different plant tissues.

The integration of anatomical, molecular, and phytopathological approaches strengthens the scientific basis necessary to advance our understanding of the so-called "anomaly" in soybeans and, above all, to offer practical answers to the production sector.

By Heloiza Leonardi (Esalq); Carlos Guilherme Theodoro e Gustavo Farias (UFMT); John Paul Rodrigues Marques (FZEA/USP); Solange Maria Bonaldo (UFMT)

Article published in issue 311 of Cultivar Grandes Culturas Magazine