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Conventional and combinatorial chlorophyll fluorescence imaging of tobamovirus-infected plants

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Photosynthetica

Abstract

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.

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Abbreviations

AS:

asymptomatic

CCD:

charge-coupled device

Chl:

chlorophyll

Chl-F:

chlorophyll fluorescence

Chl-FI:

chlorophyll fluorescence imaging

dpi:

days post-inoculation

F0 :

minimum chlorophyll fluorescence in the dark adapted state

FM :

maximum chlorophyll fluorescence in the dark adapted state

FM’:

maximum chlorophyll fluorescence in the light adapted state

FM’’:

maximum chlorophyll fluorescence after switching off the “actinic light” (protocol 2)

FP :

maximum chlorophyll fluorescence reached at the induction curve without saturating flashes

Ft :

chlorophyll fluorescence value prior to the saturating pulse in the light-adapted state

FV :

variable fluorescence

HI:

high irradiance

LDA:

Linear Discriminant Analysis

LED:

light-emitting diode

LI:

low irradiance

PAR:

photosynthetically active radiation

NPQ:

non-photochemical quenching

NPQt :

non-photochemical quenching at referred kinetics times, t

P N :

net photosynthetic rate

PMMoV-I and-S:

Italian and Spanish strains of the Pepper mild mottle virus

PS:

photosystem

R:

image resulting from the combinatorial imaging

Re:

recovered

S:

symptomatic

SFFS:

Sequential Forward Floating Search

ΦPS2 :

photosystem 2 quantum yield

ΦPS2-t :

photosystem 2 quantum yield at referred kinetics times, t [s]

References

  • Aldea, M., Hamilton, J.G., Resti, J.P., Zangerl, A.R., Berenbaum, M.R., Frank, T.D., de Lucia, E.H.: Comparison of photosynthetic damage from arthropod herbivory and pathogen infection in understory hardwood saplings.-Oecologia 149: 221–232, 2006.

    Article  PubMed  Google Scholar 

  • Allen, J.F., Mullineux, C.W.: Probing the mechanism of state transitions in oxygenic photosynthesis by chlorophyll fluorescence spectroscopy, kinetics and imaging.-In: Papageorgiou, C.G., Govindjee (ed.): Chlorophyll a Fluorescence: A Signature of Photosynthesis. Pp. 447–461. Springer, Dordrecht 2004.

    Google Scholar 

  • Alonso, E., García-Luque, I., Avila-Rincón, M.J., Wicke, B., Serra, M.T., Díaz-Ruiz, J.R.: A tobamovirus causing heavy losses in protected pepper crops in Spain.-J. Phytopathology 125: 67–76, 1989.

    Article  Google Scholar 

  • Balachandran, S., Hurry, V.M., Kelley, S.E., Osmond, C.B., Robinson, S.A., Rohozinski, J., Seaton, G.G.R., Sims, D.A.: Concepts of plant biotic stress. Some insights into the stress physiology of virus-infected plants, from the perspective of photosynthesis.-Physiol. Plant. 100: 203–213, 1997.

    Article  CAS  Google Scholar 

  • Balachandran, S., Osmond, B.C.: Susceptibility of tobacco leaves to photoinhibition following infection with two strains of tobacco mosaic virus under different light and nitrogen nutrition regimes.-Plant Physiol. 104: 1051–1057, 1994.

    PubMed  CAS  Google Scholar 

  • Balachandran, S., Osmond, B.C., Daley, F.P.: Diagnosis of the earliest strain-specific interactions between tobacco mosaic virus and chloroplasts of tobacco leaves in vivo by means of chlorophyll fluorescence imaging.-Plant Physiol. 104: 1059–1065, 1994.

    PubMed  CAS  Google Scholar 

  • Barón, M., Rahoutei, J., Lázaro, J.J., García-Luque, I.: PSII response to biotic and abiotic stress.-In: Mathis, P. (ed.): Photosynthesis: From Light to Biosphere. Vol. 4. Pp. 897–900. Kluwer Academic Publ., Dordrecht-Boston-London 1995.

    Google Scholar 

  • Berger, S., Benediktyová, Z., Matouš, K., Bonfig, K., Mueller, M.J., Nedbal, L., Roitsch, T.: Visualization of dynamics of plant-pathogen interaction by novel combination of chlorophyll fluorescence imaging and statistical analysis: Differential effects of virulent and avirulent strains of P. syringae and of oxylipins on A. thaliana.-J. exp. Bot. 58: 797–806, 2007.

    Article  PubMed  CAS  Google Scholar 

  • Berger, S., Papadopoulos, M., Schreiber, U., Kaiser, W., Roitsch, T.: Complex regulation of gene expression, photosynthesis and sugar levels by pathogen infection in tomato.-Physiol. Plant. 122: 419–428, 2004.

    Article  CAS  Google Scholar 

  • Bilger, W., Björkman, O.: Role of the xanthophyll cycle in photoprotection elucidated by measurements of light-induced absorbance changes, fluorescence and photosynthesis in leaves of Hedera canariensis.-Photosynth. Res. 25: 173–185, 1990.

    Article  CAS  Google Scholar 

  • Bonfig, K.B., Schreiber, U., Gabler, A., Roitsch, T., Berger, S.: Infection with virulent and avirulent P. syringae strains differentially effects photosynthesis and sink metabolism in Arabidopsis leaves.-Planta 225: 1–12, 2006.

    Article  PubMed  CAS  Google Scholar 

  • Chaerle, L., Hagenbeek, D., De Bruyne, E., Valcke, R., Van Der Straeten, D.: Thermal and chlorophyll-fluorescence imaging distinguish plant pathogen interaction at an early stage.-Plant Cell Physiol. 45: 887–896, 2004.

    Article  PubMed  CAS  Google Scholar 

  • Chaerle, L., Pineda, M., Romero-Aranda, R., Van Der Straeten, D., Barón, M.: Robotized thermal and chlorophyll-fluorescence imaging of Pepper mild mottle virus infection in Nicotiana benthamiana.-Plant Cell Physiol. 47: 1323–1336, 2006.

    Article  PubMed  CAS  Google Scholar 

  • Chou, H.M., Bundock, N., Rolfe, A.S., Scholes, D.J.: Infection of Arabidopsis thaliana leaves with Albugo candida (white blister rust) causes a reprogramming of host metabolism.-Mol. Plant Pathol. 2: 99–113, 2000.

    Article  Google Scholar 

  • Critchley, C., Russell, A.W.: Photoinhibition of photosynthesis in vivo: The role of protein turnover in photosystem II.-Physiol. Plant. 92: 188–196, 1994.

    Article  CAS  Google Scholar 

  • Fukunaga, K.: Introduction to Statistical Pattern Recognition. 2nd Ed.-Academic Press, New York 1990.

    Google Scholar 

  • García-Luque, I., Ferrero, M.L., Rodríguez, J.M., Alonso, E., de la Cruz, A., Sanz, A.I., Vaquero, C., Serra, M.T., Díaz-Ruíz, J.R.: The nucleotide sequence of the coat protein genes and 3’ non-coding regions of two resistance-breaking tobamoviruses in pepper shows that they are different viruses.-Arch. Virol. 131: 75–88, 1993.

    Article  PubMed  Google Scholar 

  • Genty, B., Briantais, J.-M., Baker, N.R.: The relationship between quantum yield of photosynthetic electron transport and quenching of chlorophyll fluorescence.-Biochim. biophys. Acta 990: 87–92, 1989.

    CAS  Google Scholar 

  • Horton, P., Wentworth, M., Ruban, A.: Control of the light harvesting function of chloroplast membranes: The LHCII-aggregation model for non-photochemical quenching.-FEBS Lett. 579: 4201–4206, 2005.

    Article  PubMed  CAS  Google Scholar 

  • Ireland, C.R., Long, S.P., Baker, N.R.: The relationship between carbon dioxide fixation and chlorophyll fluorescence during induction of photosynthesis in maize leaves at different temperatures and carbon dioxide concentrations.-Planta 160: 550–558, 1984.

    Article  CAS  Google Scholar 

  • Kautsky, H., Hirsch, A.: Neue Versuche zur Kohlensäureassimilation.-Naturwissenschaften 48: 964–964, 1931.

    Article  Google Scholar 

  • Kitajima, H., Butler, W.L.: Quenching of chlorophyll fluorescence and primary photochemistry in chloroplasts by dibromothymoquinone.-Biochim. biophys. Acta 376: 105–115, 1975.

    Article  PubMed  CAS  Google Scholar 

  • Lamb, E.M., Adkins, S.E., Shuler, K.D., Roberts, P.D.: Pepper mild mottle virus.-Gainesville FL: EDIS, HS-808, 2001.

    Google Scholar 

  • Lichtenthaler, H.K., Babani, F.: Light adaptation and senescence of the photosynthetic apparatus. Changes in pigment composition, chlorophyll fluorescence parameters and photosynthetic activity.-In: Papageorgiou, G.C., Govindjee (ed.): Chlorophyll a Fluorescence: A Signature of Photosynthesis. Pp. 713–736. Springer, Dordrecht 2004.

    Google Scholar 

  • Lichtenthaler, H.K., Miehé, J.A.: Fluorescence imaging as a diagnostic tool for plant stress.-Trends Plant Sci. 2: 316–320, 1997.

    Article  Google Scholar 

  • Lichtenthaler, H.K., Rinderle, U.: The role of chlorophyll fluorescence in the detection of stress conditions in plants.-CRC crit. Rev. anal. Chem. 19: S29–S85, 1988.

    Google Scholar 

  • Lohaus, G., Heldt, H.W., Osmond, C.B.: Infection with phloem limited Abutilon mosaic virus causes localized carbohydrate accumulation in leaves of Abutilon striatum: relationships to symptom development and effects on chlorophyll fluorescence quenching during photosynthetic induction.-Plant Biol. 2: 161–167, 2000.

    Article  CAS  Google Scholar 

  • Matouš, K., Benediktyová, Z., Berger, S., Roitsch, T., Nedbal, L.: Case study of combinatorial imaging: What protocol and what chlorophyll fluorescence image to use when visualizing infection of Arabidopsis thaliana by Pseudomonas syringae?-Photosynth. Res. 90: 243–253, 2006.

    Article  PubMed  CAS  Google Scholar 

  • Maxwell, K., Johnson, G.N.: Chlorophyll fluorescence — a practical guide.-J. exp. Bot. 51: 659–668, 2000.

    Article  PubMed  CAS  Google Scholar 

  • Nedbal, L., Březina, V.: Complex metabolic oscillations in plants forced by harmonic irradiance.-Biophys. J. 83: 2180–2189, 2002.

    PubMed  CAS  Google Scholar 

  • Nedbal, L., Březina, V., Adamec, F., Štys, D., Oja, V., Laisk, A., Govindjee: Negative feedback regulation is responsible for the non-linear modulation of photosynthetic activity in plants and cyanobacteria exposed to a dynamic light environment.-Biochim. biophys. Acta 1607: 5–17, 2003.

    Article  PubMed  CAS  Google Scholar 

  • Nedbal, L., Soukupová, J., Kaftan, D., Whitmarsh, J., Trtílek, M.: Kinetic imaging of chlorophyll fluorescence using modulated light.-Photosynth. Res. 66: 3–12, 2000.

    Article  PubMed  CAS  Google Scholar 

  • Nedbal, L., Whitmarsh, J.: Chlorophyll fluorescence imaging of leaves and fruits.-In: Papageorgiou, C.G., Govindjee (ed.): Chlorophyll a Fluorescence: A Signature of Photosynthesis. Pp. 389–407. Springer, Dordrecht 2004.

    Google Scholar 

  • Osmond, C.B., Daley, P.F., Badger, M.R., Lüttge, U.: Chlorophyll fluorescence quenching during photosynthetic induction in leaves of Abutilon striatum Dicks infected with Abutilon mosaic virus observed with a field-portable imaging system.-Bot. Acta 111: 390–397, 1998.

    CAS  Google Scholar 

  • Pérez-Bueno, M.L., Ciscato, M., vandeVen, M., García-Luque, I., Valcke, R., Barón, M.: Imaging viral infection: studies on Nicotiana benthamiana plants infected with the pepper mild mottle tobamovirus.-Photosynth. Res. 90: 111–123, 2006.

    Article  PubMed  CAS  Google Scholar 

  • Pérez-Bueno, M.L., Rahoutei, J., Sajnani, C., García-Luque, I., Barón, M.: Proteomic analysis of the oxygen-evolving complex of photosystem II under biotic stress. Studies on Nicotiana benthamiana infected with tobamoviruses.-Proteomics 4: 418–425, 2004.

    Article  PubMed  CAS  Google Scholar 

  • Pudil, P., Novovicova, J., Kittler, J.: Floating search methods in feature selection.-Pattern Recognition Lett. 15: 1119–1125, 1994.

    Article  Google Scholar 

  • Rahoutei, J., Barón, M., García-Luque, I., Droppa, M., Neményi, A., Horvath, G.: Effect of tobamovirus infection on the thermoluminiscence characteristics of chloroplast from infected plants.-Z. Naturforsch. 54c: 634–639, 1999.

    Google Scholar 

  • Rahoutei, J., García-Luque, I., Barón, M.: Inhibition of photosynthesis by viral infection: Effect on PSII structure and function.-Physiol. Plant. 110: 286–292, 2000.

    Article  CAS  Google Scholar 

  • Rahoutei, J., García-Luque, I., Cremona, V., Barón, M.: Effect of tobamovirus infection on PSII complex of infected plants.-In: Garab, G. (ed.): Photosynthesis: From Light to Biosphere. Vol. IV. Pp. 2761–2764. Kluwer Academic Publ., Dordrecht-Boston-London 1998.

    Google Scholar 

  • Roháček, K., Barták, M.: Technique of the modulated chlorophyll fluorescence: basic concepts, useful parameters, and some applications.-Photosynthetica 37: 339–363, 1999.

    Article  Google Scholar 

  • Ruban, A.V., Horton, P.: Regulation of non-photochemical quenching of chlorophyll fluorescence in plants.-Aust. J. Plant Physiol. 22: 221–230, 1995.

    Article  CAS  Google Scholar 

  • Sajnani, C., Zurita, J.L., Doncel, M., Ortega, J.M., Barón, M., Ducruet, J.-M.: Changes in photosynthetic metabolism induced by tobamovirus infection in Nicotiana benthamiana studied in vivo by chlorophyll thermoluminescence.-New Phytol. 175: 120–130, 2007.

    Article  PubMed  CAS  Google Scholar 

  • Scholes, J.D., Rolfe, S.A.: Photosynthesis in localised regions of oat leaves infected with crown rust (Puccinia coronata): quantitative imaging of chlorophyll fluorescence.-Planta 199: 573–582, 1996.

    Article  CAS  Google Scholar 

  • Schreiber, U., Schliwa, U., Bilger, W.: Continuous recording of photochemical and non-photochemical chlorophyll fluorescence quenching with a new type of modulation fluorometer.-Photosynth. Res. 10: 51–62, 1986.

    Article  CAS  Google Scholar 

  • Soukupová, J., Smatanová, S., Nedbal, L., Jegorov, A.: Plant response to destruxins visualized by imaging of chlorophyll fluorescence.-Physiol. Plant. 118: 399–405, 2003.

    Article  Google Scholar 

  • Swarbrick, P.J., Schulze-Lefert, P., Scholes, J.D.: Metabolic consequences of susceptibility and resistance (race-specific and broad-spectrum) in barley leaves challenged with powdery mildew.-Plant Cell Environ. 29: 1061–1076, 2006.

    Article  PubMed  CAS  Google Scholar 

  • Tang, J.Y., Zielinski, R.E., Zangerl, A.R., Crofts, A.R., Berenbaum, M.R., de Lucia, E.H.: The differential effects of herbivory by first and fourth instars of Trichoplusia ni (Lepidoptera: Noctuidae) on photosynthesis in Arabidopsis thaliana.-J. exp. Bot. 57: 527–536, 2006.

    Article  PubMed  CAS  Google Scholar 

  • van Wijk, K.J., Schnettger, B., Graf, H., Krause, G.H.: Photoinhibition and recovery in relation to heterogeneity of photosystem II.-Biochim. biophys. Acta 1142: 59–68, 1993.

    Article  Google Scholar 

  • Wetter, C., Conti, M., Alschuh, D., Tabillion, R., van Regemortel, M.H.V.: Pepper mild mottle virus, a tobamovirus infecting pepper cultivars in Sicily.-Phytopathology 74: 405–410, 1984.

    Article  Google Scholar 

  • Zangerl, A.R, Hamilton, J.G., Miller, T.J., Crofts, A.R., Oxborough, K., Berenbaum, M.R., de Lucia, E.H.: Impact of folivory on photosynthesis is greater than the sum of its holes.-Proc. nat. Acad. Sci. USA 22: 1088–1091, 2002.

    Article  CAS  Google Scholar 

  • Zhou, Y.H., Peng, Y.H., Lei, J.L., Zou, L.Y., Zheng, J.H., Yu, J.Q.: Effects of potato virus YNTN infection on gas exchange and photosystem 2 function in leaves of Solanum tuberosum L.-Photosynthetica 42: 417–423, 2004.

    Article  CAS  Google Scholar 

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Authors and Affiliations

  1. Department of Biochemistry, Molecular and Cell Biology of Plants, Estación Experimental del Zaidín, Consejo Superior de Investigaciones Científicas (CSIC), C/Profesor Albareda, n° 1, C.P. 18008, Granada, Spain

    M. Pineda & M. Barón

  2. Institute of Systems Biology and Ecology, Academy of Sciences of the Czech Republic, Zámek 136, Nové Hrady, 37333, Czech Republic

    J. Soukupová, K. Matouš & L. Nedbal

  3. Institute of Physical Biology, University of South Bohemia, Zámek 136, Nové Hrady, 37333, Czech Republic

    J. Soukupová, K. Matouš & L. Nedbal

Authors
  1. M. Pineda
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  2. J. Soukupová
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  3. K. Matouš
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  4. L. Nedbal
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  5. M. Barón
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Correspondence to M. Barón.

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Pineda, M., Soukupová, J., Matouš, K. et al. Conventional and combinatorial chlorophyll fluorescence imaging of tobamovirus-infected plants. Photosynthetica 46, 441–451 (2008). https://doi.org/10.1007/s11099-008-0076-y

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  • DOI: https://doi.org/10.1007/s11099-008-0076-y

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