The information is prepared on the basis of data from the information-analytical system RSF, informative part is represented in the author's edition. All rights belong to the authors, the use or reprinting of materials is permitted only with the prior consent of the authors.

Project titleEarly remote diagnosis of stress photosynthetic response of agricultural plants under the influence of adverse factors

KeywordsNPQ, photochemical reflectance index, remote plant status diagnostics, xanthophyll cycle, adverse factors, plant stress response, photosynthesis

Annotation
The general scientific problem that the Project 2017 was aimed at was the development of an early remote diagnostics method for the stress response of agricultural plants under the influence of adverse factors, which is based on measuring the photochemical reflectance index (PRI), and the creation of a technical system for its implementation. The high importance of this investigation was associated with the key role of systems for the optical assessment of plant physiological state for their remote monitoring and early detection of stress factors, as well as for rapid assessment of plant conditions in high-performance phenotyping. Moreover, the choice of PRI, which is calculated on the basis of two spectral bands of reflected light (531 and 570 nm), is due to its high sensitivity to changes in the state of the plant; first of all, to changes in photosynthetic processes. However, the use of PRI also has its difficulties associated with a relatively small magnitude of changes, their dependence on several processes with different characteristic times, sensitivity to measurement conditions, etc. As part of the project in 2017, we minimized some of the problems noted. In particular, the development and implementation of the method of active illumination of the studied plants with pulses of yellow-green measuring light in the PRI-imaging system reduced the distorting effect of background lighting. Measurement of the light-induced changes in PRI made it possible to reduce the influence of different initial levels of the photochemical reflectance index in different plants, and also provided the basis for a more detailed assessment of photosynthesis-related processes in the laboratory (for example, during phenotyping). The estimation of spatial heterogeneity in distribution of the photochemical reflectance index, which was based on the standard deviation of the PRI calculated for the zones of the studied plants (for peas), provided an effective indicator for monitoring their stress changes during the development of soil drought in open and protected ground. On basis of results of the Project 2017 project, the new scientific problems for Project 2020 were proposed. The solution of the following scientific problems is expected: 1. A study of the prospects for using the standard deviation of PRI in areas of vegetation cover (possibly in combination with its absolute value) to identify stress changes in agricultural plants. The statement of this problem directly follows from the results of the Project 2017, which showed that an increase in the standard deviation of PRI (m(PRI)) in the leaf or vegetation cover areas can be used to detect stress changes during soil drought in a model plant - pea. Such results open up the potential for applying a fundamentally new indicator of stress changes in plants, based on measurements of the standard deviation of PRI. This can significantly influence the global practice of using the photochemical reflection index in remote monitoring. However, testing this possibility, at a minimum, requires (i) identifying similar effects during the development of drought in other crops (e.g. wheat) and (ii) assessing the presence or absence of similar changes under the influence of other stressors of a similar nature (e.g. salinity, which also leads to water deficiency). In addition, an effect that was previously shown in the framework of the Project 2017 requires further experimental research: an increase in the standard deviation between different parts of the vegetation cover when measuring control and drought-influenced plants together. If this effect is confirmed, then it can be used to identify the boundaries of the action of stress factors on plants. 2. The study of the possibility of using standard deviation or coefficient of variation for the intensity of reflected light, measured in areas of vegetation cover at wavelengths of 531 and(or) 570 nm, to detect stress changes in plants. A prerequisite for the statement of this problem is also the increase in the standard deviation of PRI in peas during drought conditions revealed during the implementation of the Project 2017. Such an increase may be due to an increase in the standard deviation of the reflected light intensity through separate spectral channels (measuring at 531 nm or reference at 570 nm), which develops under the influence of a stressor. If the proposed hypothesis is confirmed during an experimental verification, the result will be of fundamental importance for the development of methods for remote monitoring of plant state. It will become the basis for detecting stress changes in plants based on the spatial variability of the reflected light parameters in only one narrow-band spectral channel in the yellow-green region of the spectrum. This will also create the prerequisites for the technical development of a new group of simple systems for imaging stress changes in agricultural plants, which will measure the image in one narrow spectral band and calculate the standard deviation and(or) the coefficient of variation of the reflected light intensity. 3. Development of a new modification of the PRI-imaging system using only one monochrome camera and a system for automated change of narrow-band light filters during measurement. The statement of this problem is due to the need for further development of the PRI-imaging system developed under the Project 2017. Among the technical limitations of the developed system can be noted: (i) The relative high complexity of image acquisition in the developed system. (ii) The inability to simultaneously measure several variants of PRI or other reflectance indices with a similar calculation principle. (iii) The inability to simply change the set of spectral filters used for the study. Potentially, a promising way to solve this problem can be associated with the use of a turret, which provides an automated change of filters, and one monochrome camera. The system of illumination of an object with millisecond pulses of measuring light can be left essentially unchanged; however, it is possible to replace yellow-green light with broadband white light. This will significantly expand the range of spectral indices that can be simultaneously measured using active illumination. It is important, that the specific set of spectral indices can also be formed by customers (on basis simple exchange spectral filters). At the same time, this approach may have a number of its own limitations associated with the rate of change of spectral filters in the analysis. As a result, the effectiveness of the new system modification should be compared with the effectiveness of the developed PRI-imaging system.

Expected results
The implementation of the Project 2020 is aimed at solving three large-scale tasks, on the basis of which, as expected, three main groups of results will be obtained: 1. The revealing of efficiency of applying standard deviation measurements of PRI (possibly in combination with its absolute value) at the level of the vegetation cover and its zones to detect stress changes in key agricultural plants (mainly, wheat) under the influence of economically significant stressors (for example, drought and salinity). Achieving this result opens up prospects for using a fundamentally new indicator of stress changes in plants, based on measurements of the standard deviation of PRI. This parameter can be used separately or in combination with other widely-used parameters of PRI (absolute values, light-induced changes). It can be expected that application of this combinations increase sensitivity and accuracy of revealing of plant stress responses. 2. Identification of the possibility of using a standard deviation or coefficient of variation in the intensity of reflected light, measured in areas of vegetation cover at wavelengths of 531 or 570 nm, for detecting stress changes in plants. Potentially, such a result can be of fundamental importance for the development of methods for remote monitoring of state of plant, as it will create the basis for detecting stress changes based on analysis of image, which is measured in single spectral channel. Measurements based on this method will require application of the simplest measuring system, which includes single monochromic camera and single narrowband spectral filter; additionally, the system can be equipped by module for active illumination of plants by measuring light. 3. Development of a new modification of the PRI-imaging system with the possibility of simultaneous measurement of a number of spectral indices (modified PRI, NDVI, possibly WI (or difference indexes close to WI) and others), as well as with the ability to easily configure the set of studied indices by the user. Achieving this result will provide an important tool for a comprehensive assessment of the state of plants based on reflectance indices. In particular, simultaneous measurements of typical and modified PRIs (they show different components of photosynthetic changes induced by stressors), NDVI (it show quantity of photosynthetic biomass), and, probably, WI (it show water content) can be basis of revealing of stress changes with different time scales in investigated plants. The revealing will be important for the remote sensing and high-performance phenotyping for plant breeding and zoning. In general, it can be expected that results of the Project 2020 will correspond to leading scientific research in the investigated field. They will strongly extend our tools for the remote sensing of agricultural plants and for the high-performance phenotyping. These tools will include new methods of estimation of the plant physiological state, based on measurements of spatial heterogeneity of PRI and intensities of the reflected light at 531 and 570 nm (standard deviation or variation coefficient), as well as the modified system for PRI-imaging with extended functionality. It is important, that these tools can be also used for sensing of the plant state under natural conditions; i.e. they can be potentially used for ecological monitoring.

Annotation of the results obtained in 2021
At the last stage of investigations, we solved three general tasks of the Project. (1) The efficiency of the standard deviation of the spatial distribution of a typical photochemical reflectance index was estimated on wheat under drought and on pea under salt stress. The plants cultivation and PRI measuring were performed under open field conditions. (2) The efficiency of the coefficient of variation of the spatial distribution of reflectance at 530 nm (measuring wavelength of PRI) for revealing plant stress changes was estimated under stress conditions. (3) A modified PRI-imaging system was developed. The efficiency of new system was compared with the original PRI-imaging system (Project-2017). Moreover, a number of additional tasks were resolved. Firstly, we showed that the standard deviation of the spatial distribution of the typical photochemical reflectance index increased in all explored variants (including the influence of drought on wheat and salinity on pea). It can be also noted that the changes in the standard deviation observed earlier in comparison to the changes in the absolute values of PRI. This result showed that the standard deviation could be more effective than absolute value of PRI. Besides the relationship between standard deviations and the quantum yield of photosystem II could vary significantly (from the very high to medium relationship under drought and salt stress, respectively). Such variability could limit the using the standard deviation for estimation of the of photosynthetic stress changes. However, this result did not limit application of standard deviation for detection of influence of stressors. Summary, our results confirmed the efficiency of the standard deviation of the PRI spatial distribution for remote sensing of plant stress responses. Additionally, we verified hypothesis about a significant increase in the standard deviation between the average values of PRI in individual plant areas at the border of zone of the stressor action. Experimental studies did not confirm this hypothesis because the increase of the standard deviation was weak. Secondly, we showed that the coefficient of variation of the spatial distribution reflectance at 530 nm increased in all the studied variants (the influence of drought on wheat and influence of salinity on pea). This increase always started earlier than typical PRI changes and often earlier than the standard deviation of the spatial distribution of the PRI (influence of drought on wheat). We also showed strong negative correlation between the coefficients of variation and the quantum yield of photosystem II. Summary, our result confirmed the perceptivity of the coefficient of variation of the spatial distribution of reflectance at 530 nm for detection of influence of stressors on plants and possible estimation of photosynthetic parameters. Thirdly, a modified PRI-imaging system was developed for registration of reflectance at several spectral bands (including 448 nm, 510 nm, 532 nm, 546 nm, 572 nm and 685 nm). This system permits to measure the typical photochemical reflectance index (based on 532 and 572 nm), modified photochemical reflectance indices (based on 510 and 572 nm and on 546 and 572 nm) and additional differential reflectance indices (based on 448 and 572 nm and on 685 and 572 nm). Comparison of the developed modified PRI-imaging system (Project-2020) with the previous system (Project-2017) showed similar efficiency of both systems for measuring of typical PRIs and their standard deviations. However, the measurements of PRI by modified system were more accurate; and the previous system was more effective for estimation of changes in standard deviations. Besides, the using of the modified PRI imaging system revealed the influence of drought on the modified PRI and additional difference indices in pea and on the standard deviations of the spatial distribution of these indices and the coefficients of variation of reflectance in the investigated spectral bands. The last result is preliminary; however, it is perspective for the development of new methods for remote sensing of stress changes in plants. Additionally, we analyzed efficiency of new reflectance indices. These indices showed high sensitivity to the influence of stressors on plants. We identified different directions of changes in modified PRI under stress conditions. The theoretical analysis showed increase in the heterogeneity of the electrophysiological parameters of cells and, as a result, a possible increase in the heterogeneity of the processes of photosynthesis and reflectance under the action of stressors.

Publications

1. Sukhova E., Kior D., Kior A., Yudina L., Zolin Y., Gromova E., Sukhov, V. New Normalized Difference Reflectance Indices for Estimation of Soil Drought Influence on Pea and Wheat Remote Sensing, V. 14, N. 7, Article 1731 (year - 2022) https://doi.org/10.3390/rs14071731

2. Sukhova E., Ratnitsyna D., Sukhov V. Stochastic Spatial Heterogeneity in Activities of H+-ATP-Ases in Electrically Connected Plant Cells Decreases Threshold for Cooling-Induced Electrical Responses International Journal of Molecular Sciences, V. 22, N. 15, Article 8254 (year - 2021) https://doi.org/10.3390/ijms22158254

3. Sukhova E., Yudina L., Kior A., Kior D., Popova A., Zolin Y., Gromova E., Sukhov V. Modified Photochemical Reflectance Indices as New Tool for Revealing Influence of Drought and Heat on Pea and Wheat Plants Plants, - (year - 2022)

4. Ratnitsyna D., Sukhova E., Sukhov V. Разработка математической модели фотосинтеза на основе модели Farquhar-von Caemmerer-Berry, включающей в себя световое повреждение фотосинтетического аппарата Биосистемы: организация, поведение, управление: Тезисы докладов 75-й Всероссийской с международным участием школыконференции молодых ученых (Н. Новгород, 19–22 апреля 2022 г.). Н. Новгород, Университет Лобачевского. 2022. 272 с., С. 190 (year - 2022)

5. Sukhov V., Sukhova E. Вызванные локальным действием абиотических стрессоров системные фотосинтетические ответы растений: механизмы и дистанционный мониторинг IX Съезд Российского фотобиологического общества. Всероссийская конференция "Современные проблемы фотобиологии". Материалы съезда. Пущино, 2021. 182 стр., С. 67 (year - 2021)

6. Yudina L., Sukhova E., Sukhov V. Оценка фотохимического индекса отражения растений при тепловом стрессе и почвенной засухе IX Съезд Российского фотобиологического общества. Всероссийская конференция "Современные проблемы фотобиологии". Материалы съезда. Пущино, 2021. 182 стр., С. 173 (year - 2021)

7. Zolin Y., Kior A., Sukhova E., Sukhov V. Параметры пространственного распределения фотохимического индекса отражения и отражения на длине волны 531 нм показывают развитие стрессовых изменений растений при действии засухи и засоления Биосистемы: организация, поведение, управление: Тезисы докладов 75-й Всероссийской с международным участием школыконференции молодых ученых (Н. Новгород, 19–22 апреля 2022 г.). Н. Новгород, Университет Лобачевского. 2022. 272 с., С. 82 (year - 2022)

8. - Система для измерения фотохимического индекса отражения PRI у растений -, 2746690 (year - )

9. - Осторожно, двери открываются. Десять российских научных исследований 2021 года, которыми не стыдно хвастаться Коммерсантъ, Наука в России. 30.12.2021, 21:11 (year - )

Annotation of the results obtained in 2020
The Project is devoted to developing methods of plant remote sensing (early revealing actions of stressors) based on measuring the photochemical reflectance index. The following results were shown in the current stage of the Project: 1. Hypothesis about increasing spatial heterogeneity in distribution of PRI measured in a plant canopy and decreasing absolute value of the index under actions of stressors was supported. Investigations were performed on wheat and pea plants in controlled conditions of growth. The results were shown in investigations of soil drought (wheat) and salt stress (pea). 2. It was revealed that increasing the standard deviation between averaged PRI in ROIs can be used as estimator of frontier between zone with stressor action and zone without the action. The result was shown on basis of simultaneous analysis of control and experimental plants, which were acted by stressors, in controlled conditions of growth. It should be noted that the effect is only observed at strong changes in absolute value of PRI under actions of stressors. In particular, the effect was observed at action of soil drought on wheat but it was absent at salt stress in pea. 3. It was revealed that increasing spatial heterogeneity of intensity of reflected light at 531 nm, which was estimated on basis of variation coefficients of the intensity, can be used as estimator of actions of stressors on plants. The result can be basis of future developing a simple method of estimation of plant stress changes with using imaging at single spectral channel only. The efficiency of the coefficient variation using in comparison to measurements of parameters of PRI can be vary. In some cases, the efficiency is higher than efficiency of using PRI parameters (e.g., at salt stress-induced changes in peas). 4. In collaboration with I.V. Turchin’s scientific group (Institute of Applied Physics, LLC “BFV”), we proposed principle scheme of a new system of multispectral PRI imaging. The scheme includes light source (LED) for active plant illumination, digital camera with lens for image registration, tool for automatic replace of narrowband spectral filters (including the filters with specific spectral bands), controller on basis of ARM for control of the system, current and power sources. Characteristics of the components and there types were selected. Manufacturing of the system was initiated. 5. Additional investigations, which were based on the complex analysis of stressors-induced changes in all possible spectral indices in 400-700 nm spectral range, showed the new reflectance indices, which are sensitive to early changes in plants under soil drought. Also, we developed heat maps of correlation coefficients between the reflectance indices in 400-700 nm spectral ranges and photosynthetic parameters at the systemic plant response induced by local damages. The results can be basis for a search of additional spectral indices, which can be measured with using the new system of multispectral PRI imaging.

Publications

1. Sukhov V., Sukhova E., Khlopkov A., Yudina L., Ryabkova A., Telnykh A., Sergeeva E., Vodeneev V., Turchin I. Proximal imaging of changes in photochemical reflectance index in leaves based on using pulses of green-yellow light Remote Sensing, V. 13, N. 9, Article 1762 (year - 2021) https://doi.org/10.3390/rs13091762

2. Sukhova E., Yudina L., Gromova E., Ryabkova A., Vodeneev V., Sukhov V. Influence of local burning on difference reflectance indices based on 400–700 nm wavelengths in leaves of pea seedlings Plants, V. 10, N. 5, Article 876. (year - 2021) https://doi.org/10.3390/plants10050878

3. Sukhova E., Yudina L., Gromova E., Ryabkova A.Y, Kior D., Sukhov V. Complex analysis of the efficiency of difference reflectance indices on the basis of 400–700 nm wavelengths for revealing the influences of water shortage and heating on plant seedlings Remote Sensing, V. 13. N. 5, Article 962. (year - 2021) https://doi.org/10.3390/rs13050962

4. Ryabkova A.Yu., Sukhova E.M., Sukhov V.S. Разработка методов раннего выявления действия засухи на основе измерения отраженного света (531 нм и 570 нм). Биосистемы: организация, поведение, управление: Тезисы докладов 74-й Всероссийской с международным участием школы конференции молодых ученых. Н.Новгород, Университет Лобачевского. 2021., 261 с. (year - 2021)

5. Sukhova E.M., Sukhov V.S. Фотохимический индекс отражения: применение для анализа эффективности фотосинтеза и перспективы его применения Биосистемы: организация, поведение, управление: Тезисы докладов 73-й Всероссийской с международным участием школы конференции молодых ученых (Н.Новгород, 28–30 октября 2020 г.)., Н.Новгород, Университет Лобачевского. 2020. С. 199. (year - 2020)

6. - Растениям поставят точный диагноз Российская газета, Российская газета - Неделя - Приволжье № 214(8268) (year - )