BcXYG1, a Secreted Xyloglucanase from Botrytis cinerea, Triggers Both Cell Death and Plant Immune Responses.

In search of Botrytis cinerea cell death-inducing proteins, we found a xyloglucanase (BcXYG1) that induced strong necrosis and a resistance response in dicot plants. Expression of the BcXYG1 gene was strongly induced during the first 12 h post inoculation, and analysis of disease dynamics using PathTrack showed that a B. cinerea strain overexpressing BcXYG1 produced early local necrosis, supporting a role of BcXYG1 as an early cell death-inducing factor. The xyloglucanase activity of BcXYG1 was not necessary for the induction of necrosis and plant resistance, as a mutant of BcXYG1 lacking the xyloglucanase enzymatic activity retained both functions. Residues in two exposed loops on the surface of BcXYG1 were found to be necessary for the induction of cell death but not to induce plant resistance. Further analyses showed that BcXYG1 is apoplastic and possibly interacts with the proteins of the plant cell membrane and also that the BcXYG1 cell death-promoting signal is mediated by the leucine-rich repeat receptor-like kinases BAK1 and SOBIR1. Our findings support the role of cell death-inducing proteins in establishing the infection of necrotrophic pathogens and highlight the recognition of fungal apoplastic proteins by the plant immune system as an important mechanism of resistance against this class of pathogens.

Temporal Dynamics of Localized Exciton-Polaritons in Composite Organic-Plasmonic Metasurfaces.

We use femtosecond transient absorption spectroscopy to study the temporal dynamics of strongly coupled exciton-plasmon polaritons in metasurfaces of aluminum nanoantennas coated with J-aggregate molecules. Compared with the thermal nonlinearities of aluminum nanoantennas, the exciton-plasmon hybridization introduces strong ultrafast nonlinearities in the composite metasurfaces. Within femtoseconds after the pump excitation, the plasmonic resonance is broadened and shifted, showcasing its high sensitivity to excited-state modification of the molecular surroundings. In addition, we observe temporal oscillations due to the deep subangstrom acoustic breathing modes of the nanoantennas in both bare and hybrid metasurfaces. Finally, unlike the dynamics of hybrid states in optical microcavities, here, ground-state bleaching is observed with a significantly longer relaxation time at the upper polariton band.

Aluminum Nanoantenna Complexes for Strong Coupling between Excitons and Localized Surface Plasmons.

We study the optical dynamics in complexes of aluminum nanoantennas coated with molecular J-aggregates and find that they provide an excellent platform for the formation of hybrid exciton-localized surface plasmons. Giant Rabi splitting of 0.4 eV, which corresponds to ∼10 fs energy transfer cycle, is observed in spectral transmittance. We show that the nanoantennas can be used to manipulate the polarization of hybrid states and to confine their mode volumes. In addition, we observe enhancement of the photoluminescence due to enhanced absorption and increase in the local density of states at the exciton-localized surface plasmon energies. With recent emerging technological applications based on strongly coupled light-matter states, this study opens new possibilities to explore and utilize the unique properties of hybrid states over all of the visible region down to ultraviolet frequencies in nanoscale, technologically compatible, integrated platforms based on aluminum.

Polarization controlled coupling and shaping of surface plasmon polaritons by nanoantenna arrays.

We demonstrate experimentally the use of ordered arrays of nanoantennas for polarization controlled plasmonic beam shaping and excitation. Rod- and cross-shaped nanoantennas are used as local point-like sources of surface plasmon polaritons, and the desired phase of the generated plasmonic beam is set directly through their spatial arrangement. The polarization selectivity of the optical nanoantennas allows us to further control the excitation, enabling the realization of a variety of complex and functional plasmonic beam shaping elements. We demonstrate this concept by generating plasmonic self-accelerating beams, plasmonic bottle beams, and switchable dual-focii plasmonic lenses. The freedom in the design and arrangement of these nanoantennas enables us to specifically tailor and control the shapes, wavelengths, and coupling efficiencies of complex plasmonic beams.

Inverting singlet and triplet excited states using strong light-matter coupling.

In organic microcavities, hybrid light-matter states can form with energies that differ from the bare molecular excitation energies by nearly 1 eV. A timely question, given the recent advances in the development of thermally activated delayed fluorescence materials, is whether strong light-matter coupling can be used to invert the ordering of singlet and triplet states and, in addition, enhance reverse intersystem crossing (RISC) rates. Here, we demonstrate a complete inversion of the singlet lower polariton and triplet excited states. We also unambiguously measure the RISC rate in strongly coupled organic microcavities and find that, regardless of the large energy level shifts, it is unchanged compared to films of the bare molecules. This observation is a consequence of slow RISC to the lower polariton due to the delocalized nature of the state across many molecules and an inability to compete with RISC to the dark exciton reservoir.

Characterization of Botrytis-plant interactions using PathTrack© -an automated system for dynamic analysis of disease development.

The measurement of disease development is integral in studies on plant-microbe interactions. To address the need for a dynamic and quantitative disease evaluation, we developed PathTrack© , and used it to analyse the interaction of plants with Botrytis cinerea. PathTrack© is composed of an infection chamber, a photography unit and software that produces video files and numerical values of disease progression. We identified a previously unrecognized infection stage and determined numerical parameters of pathogenic development. Using these parameters, we identified differences in disease dynamics between seemingly similar B. cinerea pathogenicity mutants, and revealed new details on plant susceptibility to the fungus. We showed that the difference between the lesion expansion rate on leaves and colony spreading rate on artificial medium reflects the levels of the plant immune system, suggesting that this parameter can be used to quantify plant defence. Our results shed new light and reveal new details of the interaction between the model necrotrophic pathogen B. cinerea and plants. The concept that we present is universal and may be applied to facilitate the study of various types of plant-pathogen association.

Measurement of apoptosis by SCAN©, a system for counting and analysis of fluorescently labelled nuclei.

Apoptosis-like programmed cell death (A-PCD) is a universal process common to all types of eukaryotic organisms. Because A-PCD-associated processes are conserved, it is possible to define A-PCD by a standard set of markers. Many of the popular methods to measure A-PCD make use of fluorescent ligands that change in intensity or cellular localization during A-PCD. In single cell organisms, it is possible to quantify levels of A-PCD by scoring the number of apoptotic cells using flow cytometry instruments. In a multicellular organism, quantification of A-PCD is more problematic due to the complex nature of the tissue. The situation is further complicated in filamentous fungi, in which nuclei are divided between compartments, each containing a number of nuclei, which can also migrate between the compartments. We developed SCAN©, a System for Counting and Analysis of Nuclei, and used it to measure A-PCD according to two markers - chromatin condensation and DNA strand breaks. The package includes three modules designed for counting the number of nuclei in multi-nucleated domains, scoring the relative number of nuclei with condensed chromatin, and calculating the relative number of nuclei with DNA strand breaks. The method provides equal or better results compared with manual counting, the analysis is fast and can be applied on large data sets. While we demonstrated the utility of the software for measurement of A-PCD in fungi, the method is readily adopted for measurement of A-PCD in other types of multicellular specimens.