Application of chlorophyll fluorescence imaging in duckweed ecotoxicological testing
| dc.contributor.advisor | Oláh, Viktor | |
| dc.contributor.author | Irfan, Muhammad | |
| dc.contributor.department | Juhász-Nagy Pál doktori iskola | hu |
| dc.contributor.submitterdep | Természettudományi és Technológiai Kar::Biológiai és Ökológiai Intézet::Növénytani Tanszék | |
| dc.date.accessioned | 2024-08-28T06:16:15Z | |
| dc.date.available | 2024-08-28T06:16:15Z | |
| dc.date.created | 2024-08-26 | |
| dc.date.defended | 2024-10-17 | |
| dc.description.abstract | Increasing freshwater pollution due to anthropogenic activities have resulted in an immediate need for in situ water quality monitoring for management and conservation purposes. Different bioindicator organisms are being used to monitor water quality and model the toxicological effects of environmental pollutants. Using duckweeds in toxicological studies is becoming common due to their superior characteristics including fast growth rate, easy management, and faster response rate. In addition to classical growth-based toxicological assays, chlorophyll fluorescence (ChlF) induction-based methods are also becoming popular in duckweed toxicity tests as non-destructive, fast and easily applicable way to measure photosynthetic inhibition under toxicant exposure. In the growth-based toxicological assays, the adverse effects of the applied toxicants are mainly characterized as changes in the growth rates of surface area or production of new fronds. The ChlF-based methods, on the other hand, rely on the measurement of ground and maximum fluorescence levels under dark- and light-adapted states of the photosystem. These basic parameters are then used to derive many ChlF-based endpoints for the measurement of toxic effects. In this research, two duckweed species, namely Spirodela polyrhiza and Lemna gibba were used to assess the most sensitive endpoints using Pulse Amplitude Modulated (PAM) ChlF method and to possibly reduce the required measurement duration. The sensitivity of ChlF-based endpoints was then compared with the most applicable and most sensitive surface area growth-based endpoint to check the potential of ChlF alone to substitute the growth-based endpoints. Another aim of this research was to measure these endpoints from a small, less resource intensive and comparatively short-term tissue-culture plate-based system instead of the standardized duckweed testing system with higher infrastructure and resource requirements. For this purpose, plants were grown in smaller volume of test solutions for a shorter period. In short, the efforts were made to significantly reduce the time and effort required for the standard assay methods while producing higher throughput and increasing the efficiency of the tests. According to our findings with S. polyrhiza, many ChlF-endpoints responded to the applied treatments with varying degrees of sensitivity. Out of the tested ChlF-based endpoints, the sensitivity of dark-adapted chlorophyll fluorescence endpoint Fv/Fo was notably higher than that of the widely used Fv/Fm. On the other hand, Y(II) proved to be amongst the most sensitive light-adapted endpoints. The results emphasized the importance to consider reporting information on Fv/Fo as well in addition to Fv/Fm. The sensitivity of the selected highly responsive endpoints (i.e., Y(II) and Fv/Fo) were then subjected to comparative toxicity tests using different metals and metalloids in duckweed L. gibba. During these tests, the ChlF-generated images were also used to simultaneously measure the growth-based parameters of the test cultures. On the one hand, the results with L. gibba supported the fact that using the small-scale, multi-well-plate-based testing system could come in handy in terms of saving space, growth- and analysis-duration and required smaller volumes of testing solutions. On the other hand, the sensitivity of such growth inhibition tests was lower as compared to standard tests. Despite such lower sensitivity, the tested elements showed comparable order of phytotoxicity to previously conducted OECD- and ISO-conform tests. The responsiveness of individual tested endpoints, based on their calculated effective concentrations, were also noticeably different. The results of this study also supported the higher sensitivity of growth-inhibition-based endpoints and hence ruled out their complete substitution by ChlF-based phytotoxicity endpoints in duckweed toxicity testing. Another observation of this study was the underestimation of photosynthesis-inhibiting effects due to the exclusion of chlorotic frond parts when exposed to an element with high acute phytotoxic potential. The results of this study also suggested that changes in the photochemical efficiency of duckweeds were not necessarily corresponding to the growth responses due to such exclusions. Another attempt to make the measurements shorter, that is using ChlF-based Fm images to measure growth-based parameters of the test cultures, also proved to bias the measurement. This was due to chlorotic regions in fronds that appeared as virtual loss in total surface area causing an underestimation in this growth parameter. Despite the comparatively lower sensitivity and methodological constraints, ChlF-based phenotyping techniques can increase the throughput of toxicity assays by providing a non-invasive tool for physiological monitoring. Furthermore, the utilization of imaging techniques in ChlF allowed for the simultaneous measurement of duckweed growth and photosynthetic responses in phytotoxicity studies, resulting in a considerable reduction in testing duration. Finally, using the small-scale multi-well-plate-based testing system in combination with ChlF imaging technique, duckweed phytotoxicity assays can facilitate simultaneous screening of large sample series or multiple duckweed species/clones within a short duration. | |
| dc.format.extent | 112 | |
| dc.identifier.uri | https://hdl.handle.net/2437/379345 | |
| dc.language.iso | en | |
| dc.subject | Duckweed | |
| dc.subject | Chlorophyll fluorescence | |
| dc.subject | Lemna gibba | |
| dc.subject | Spirodela polyrhiza | |
| dc.subject | Heavy metals | |
| dc.subject | Toxicity | |
| dc.subject.sciencefield | Természettudományok | hu |
| dc.title | Application of chlorophyll fluorescence imaging in duckweed ecotoxicological testing | |
| dc.title.translated | Application of chlorophyll fluorescence imaging in duckweed ecotoxicological testing |
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