We compared by chlorophyll (Chl) fluorescence imaging the effects of two strains of the same virus (Italian and Spanish strains of the Pepper mild mottle virus − PMMoV-I and -S, respectively) in the host plant Nicotiana benthamiana. The infection was visualized either using conventional Chl fluorescence parameters or by an advanced statistical approach, yielding a combinatorial set of images that enhances the contrast between control and PMMoV-infected plants in the early infection steps. Among the conventional Chl fluorescence parameters, the non-photochemical quenching parameter NPQ was found to be an effective PMMoV infection reporter in asymptomatic leaves of N. benthamiana, detecting an intermediate infection phase. The combinatorial imaging revealed the infection earlier than any of the standard Chl fluorescence parameters, detecting the PMMoV-S infection as soon as 4 d post-inoculation (dpi), and PMMoV-I infection at 6 dpi; the delay correlates with the lower virulence of the last viral strain.Additional key words: biotic stress; Nicotiana benthamiana; non-photochemical quenching; Pepper mild mottle virus.

Fluorometers that measure the kinetics of chlorophyll fluorescence have become invaluable tools for determining the photosynthetic performance of plants. Many of these instruments use high frequency modulated light to measure the rate, efficiency and regulation of photosynthesis. The technique is non-invasive and is effective under diverse environmental conditions. Recently, imaging fluorometers have been introduced that reveal variability in photosynthesis over the surface of a leaf or between individual plants. Most imaging instruments depend on continuous light or low frequency modulated light for fluorescence excitation, which imposes serious limitations on measurements of the fluorescence parameters, especially the minimum fluorescence (F(0)) and variable fluorescence (F(V)). Here, we describe a new instrument that combines the advantage of high frequency modulated light with two-dimensional imaging of chlorophyll fluorescence. The fluorometer produces dynamic images of chlorophyll fluorescence from leaves or plants, providing accurate mapping of F(0) and F(V), and non-photochemical quenching. A significant feature of the instrument is that it can record fluorescence images of leaves in daylight under field conditions.

No abstract

Kinetic fluorescence imaging was used to set a new detection limit for plant exposure to low levels of destruxins – phytotoxins of Alternaria brassicae. A general experimental algorithm is presented that can be used to identify the combination of fluorescence parameters providing the highest contrast between the affected and unaffected plants or plant segments. Leaves of canola (Brassica napus) and white mustard (Sinapis alba) were exposed to various concentrations of destruxins and images of key fluorescence signals (F0, FM, FP, and of FS) were captured in a single kinetic experiment. Contrast was quantified within these images between the leaf areas exposed to destruxins and the untreated areas. The highest contrast was found in the image constructed by pixel‐to‐pixel division of images F0 by FP and F0 by FM. Using the F0/FM ratio image, we were able to detect exposure to destruxin concentration as low as approximately 0.05 mg l−1 applied to canola leaf and approximately 10 mg l−1 when applied to mustard. The detection limits were significantly lower than those obtained by optical microscopy indicating that kinetic chlorophyll fluorescence imaging can be used as a diagnostic tool in screening for varieties with an enhanced resistance to destruxins of Alternaria brassicae.

Remotely sensed passive chlorophyll fluorescence emission has a potential to become one of the major global-scale reporter signals on vegetation performance and stress. In contrast to the actively probed parameters such as maximal (FM′) or minimal (F0′) emission, the steady-state chlorophyll fluorescence, Chl-FS, (FM′ > Chl-FS > F0′) has not been adequately studied. Using fluorescence imaging of leaves, we explored the modulation of Chl-FS by actinic irradiance and by temperature in laboratory, as well as the changes that occurred in three coniferous and broadleaf plant species grown in field. The experiments revealed that Chl-FS is largely insensitive to the incident irradiance once this is above early morning or late evening levels. The characteristic, pre-noon measured Chl-FS correlated positively with the CO2 assimilation rate when measured in field during the year. It was low and stable in the cold winter months and steeply increased with the spring onset. The high values of the characteristic Chl-FS persisted throughout the vegetation season and rapidly decreased in the fall. The seasonal Chl-FS transitions coincided with the last spring frosts or the first fall frosts that persisted for several consecutive nights. The transitions were marked by an elevated variability of the Chl-FS signal. We propose that the signal variability occurring during the transition periods can be used to detect from satellites the beginning and the end of the photosynthetic activity in evergreen canopies of the temperate zone.