BACKGROUND
plants are exposed to various forms of mechanical stress caused by rain, snow, wind, animals, pathogens or plants themselves.
such mechanical stimuli induce responses in the plant that were shown in many cases to have an adaptive value  <cit> .
a classical example is the response of trees to wind that results in shorter and thicker trunks.
reaction or compression wood is an anatomical consequence of sensing mechanical stress with subsequent lignification of cell walls  <cit> .
plants also respond to a more delicate mechanical stress referred to as touch that leads to nastic or tropic responses .
classical examples include the folding of mimosa pudica’s leaflets, the leaf closure of the venus fly trap or the coiling of tendrils  <cit> .
such stimuli lead to visible responses such as a reorientation of organs or changes in shape allowing catching an insect or improved anchorage.
the response of plants to mechanical stimuli can also be more discrete without any apparent overt changes.
for example, mechanical stress associated with damage or wounds can lead to increased resistance to insects  <cit>  or fungal pathogens  <cit> .
a closer look at the response to wounding has shown that it induces biochemical and molecular changes often associated with subsequently induced resistance mechanisms.
for instance, wounded plants produce reactive oxygen species   <cit> , undergo changes in lignification  <cit> , ja, other hormones or wound signals  <cit>  and exhibit changes in gene expression  <cit>  that are associated with induced defense reactions.
in a previous study we have shown that wounding of arabidopsis thaliana leaves induces a strong and transient immunity to botrytis cinerea the causal agent of grey mould  <cit> .
the expression of genes for camalexin biosynthesis and of glutathione-s-transferase, the activity of a map kinase activity and the accumulation of camalexin are primed by wounding  <cit> .
wound-induced immunity is independent of the major plant defense pathways involving salicylic acid, ja or ethylene, but depends on glutathione  <cit> .
recently, we have shown that wounding leads to the formation of ros within minutes and ros are required for wound–induced resistance to b. cinerea <cit> .
a strong constitutive resistance to b. cinerea also takes place in mutants such as bdg and lacs <dig>  defective in the production of a functional cuticle and displaying a phenotype of enhanced cuticular permeability  <cit> .
moreover, leaf surfaces treated with cutinase produced ros and became more protected to b. cinerea <cit> .
thus, increased permeability of the cuticle is linked to ros formation and resistance to b. cinerea.
in this study, we have further explored the responses of a. thaliana such as ros and resistance to b. cinerea in leaves that are subjected to more gentle form of mechanical stimulation.
RESULTS
sms treatment of a. thaliana leaves induces ros and resistance to b. cinerea
we have treated leaves of a. thaliana by gently rubbing them between thumb and forefinger, a mechanical stress that is herewith referred to as a soft mechanical stimulation .
the inoculation of leaves immediately after sms with spores of b. cinerea led to a strong decrease in lesion size .
a rapid burst in ros evidenced by a green fluorescence was observed in leaves infiltrated with 5--carboxy- <dig> -dichlorodihydrofluorescein diacetate  immediately after sms .
dcf-da detects a broad range of oxidizing reagents including h2o <dig> and o2- and its use has been previously described  <cit> .
sms-induced resistance as well as wound-induced resistance were still detected in mutants of nadph oxidase d  and f  as well as in the double mutant  meaning that others rboh proteins are implicated in the formation of ros .
the response to sms showed a dose-dependence .
applying only one event of sms already lead to a small but detectable decrease in lesion size  as well as production of discrete patches of green dcf-da-fluorescence .
we used  <dig> stimulations throughout this study as this lead to the strongest effects.
unstimulated upper leaves of plants stimulated on one lower leaf were not protected against b. cinerea  showing the absence of a systemic effect.
moreover, wearing latex gloves was equally effective for sms-induced resistance .
the sms-induced resistance to b. cinerea was transient: when plants were inoculated 8 h after sms, about 50% of the resistance was lost and  <dig> h after sms, plants were fully susceptible .
sms was followed by a rapid change in intracellular calcium as detected using both yellow cameleon- or aequorin-expressing plants  <cit>  .
the expression of so-called touch genes previously associated with mechanical stimulation such as tch <dig>  tch <dig>  cml <dig> and cml <dig> <cit>  was also induced  <dig>  to 1 h after sms treatment .
the effect of sms could still be observed in mutants of ja biosynthesis and signaling .
the ethylene mutant ein2- <dig> also responds to sms .
sms is not accompanied by cellular damage
we next examined the occurrence of overt wounding after sms, since wounding was shown previously to strongly affect resistance to b. cinerea <cit> .
macroscopic signs of wounding were not visible on sms-treated leaves.
nevertheless we assessed the effect of sms on the cell surface by scanning electron microscopy  of live leaf surfaces as well as vital staining using trypan blue.
sms-treated epidermal cells looked perfectly turgid when viewed under the sem .
the waxy surface of some cells appeared slightly affected but the cells themselves retained turgidity.
some trichomes and cells at their base displayed damage .
when sms-treated leaves were stained using trypan blue, a vital dye that marks the presence of dead cells, epidermal cells were essentially intact 1 h after treatment with the exception of isolated cell groups that stained in blue at the basis of trichomes , in agreement with observations made using the sem directly after sms.
thus, sms did not lead to massive cellular damage compared to wounding with forceps that was observed previously  <cit> .
the question remains if the damaged cells at the basis of the trichomes might constitute a sufficiently strong wound stimulus to induce ros and resistance.
this question was approached using the trichome-less glabrous gl <dig> mutant of arabidopsis  <cit> .
sem images of leaf surfaces of the gl <dig> mutant were compared to wt plants and in both plants cells retained turgidity after sms .
no damaged cells were observed using the vital stain trypan blue in gl <dig> mutant after sms .
in fact, the untreated gl <dig> mutant is as susceptible to b. cinerea as wt plants and after sms, gl <dig> displayed resistance to b. cinerea to the same extent as the wild type .
thus, sms induced resistance independently of the presence of trichomes and sms-induced resistance to b. cinerea is not based on wounding of cells.
alterations in cuticular permeability and sms
an accumulation of ros was previously observed in a. thaliana plants displaying increased cuticular permeability such as mutants defective in cuticle biosynthesis or in the formation of abscisic acid   <cit> .
therefore we have determined if the sms-stimulated leaves underwent a change in cuticular permeability.
the permeability of the cuticle of sms-stimulated leaves was assessed using various diagnostic tests.
sms-treated leaves displayed an increase in cuticular permeability as collectively indicated by increased chlorophyll leakage, retention of toluidine blue as well as calcofluor white staining .
sms is not accompanied by changes in aba levels
wound-induced resistance to b. cinerea is lost when wounded plants are not maintained under a humid environment .
this loss is caused by aba, the level of which increases under dry conditions   <cit> .
in accordance, mutants impaired in aba were fully resistant after wounding, both when plants were maintained under humid or dry environments  <cit> .
interestingly, increase in resistance and production in ros were observed whether plants were maintained at humid or dry conditions after sms .
consequently, we also determined possible changes in the level of aba.
no changes were observed between the levels of aba after sms in plants maintained under humid or dry conditions .
this marks a clear difference between sms and wound-induced resistance.
sms and the surface wax layer
since sms likely perturbs the waxy surface of the plant without much damage to the underlying cells , we explored the importance of the wax layer on the leaf surface.
we have used the myb96- <dig> mutant affected in the transcription factor myb <dig>  involved in the biosynthesis enzymes condensing very-long-chain fatty acids involved in cuticular wax biosynthesis  <cit> .
untreated myb96- <dig> mutants displayed increased resistance to b. cinerea .
the ros response of myb96- <dig> mutants to b. cinerea was much faster than in the wild type since green fluorescence was detected already 3 hours after inoculation with b. cinerea .
the myb96- <dig> mutant also displayed a slight modification of permeability as indicated by the toluidine blue and calcofluor white test, but this modification was not detected with the chlorophyll leakage test .
thus, myb96- <dig> altered in the wax layer displays a somewhat similar syndrome  although less obvious that the wild type leaves after sms treatment.
sms and leaf diffusates
changes in the permeability of the cuticle were shown to be associated with the leakage of diffusates that prevent the development of b. cinerea in vitro and in vivo <cit> .
we have tested if bioactive diffusates can be obtained from leaf surfaces of sms-treated wt or myb96- <dig> plants.
sms applied to gl <dig> and myb96- <dig> mutants was similarly effective as on sms-treated wt plants .
without sms, diffusates collected from the surfaces of both gl <dig> and myb96- <dig> plants were inactive similarly to those from wt plants.
thus sms acted on leaf surfaces in a comparable way in plants or mutants displaying increased cuticular permeability  <cit> .
discussion
wounding inflicted by clamping leaves with forceps or puncturing with a needle induces a strong immunity of a. thaliana to b. cinerea <cit> .
in this study, we have explored the effect of softer forms of mechanical stimulation on the resistance of a. thaliana to b. cinerea.
in particular, we have observed that a gentle mechanical stimulus applied to the surface of the leaf induced a transient and localized resistance to b. cinerea.
plants are known to be equipped with a sensitive and discriminative sensory system for the detection of molecular patterns generated by pathogens   <cit> .
here we show that a gentle mechanical stress can also be perceived in a differentiated way and lead to specific plant responses that include resistance against a virulent necrotrophic fungus.
how does sms compare to wounding?
overall, the results on sms-induced resistance to b. cinerea overlap with wound-induced resistance of a. thaliana leaves  <cit> .
the results presented here further the published observations on wounding by showing that a soft mechanical friction of the surface layer without wounding the underlying cell is already enough to induce resistance.
the absence of overt cellular breakage after sms is supported by the absence of change in the levels of aba after sms under dry or humid conditions  and the observation of leaf surfaces of wt and glabrous mutants after sms .
despite this, sms-treated plants as well as the waxless mutant display an increased permeability to toluidine blue or calcofluor white, indicating that the cuticular barrier is affected to a certain extent.
the altered cuticular permeability might allow the diffusion of a bioactive molecule observed in sms-treated plants , similarly as in cuticle-defective mutants  <cit> .
both wounding and sms lead to a rapid and important release of ros.
a number of reports have associated mechanical stress with an increased production of ros  <cit> .
for example, rubbing tomato plant internodes results in a rapid and lasting accumulation of h2o <dig> <cit> .
ros are well known for their effect as intracellular signals and were shown to be involved in the activation of defenses in response to biotic and abiotic stress  <cit> .
sms-induced resistance to b. cinerea still takes place in atrbohd, atrbohf and atrbohd/f mutants of nadph oxidoreductase making it unlikely that these nadph oxidoreductases are involved .
it cannot be excluded that some other arabidopsis nadph oxidoreductases are involved.
this is similar to results obtained with these mutants after wounding  <cit> .
in addition, sms- like wound-induced resistance are both independent of ja signaling   <cit> .
how is sms perceived by the plant?
the results presented here lend themselves to a similar interpretation as the studies on resistance to b. cinerea observed in a. thaliana after wounding or in plants with defective cuticles  <cit> .
sms might modify the plant surface making it more permeable thus allowing a better transfer of damps  or mamps  through the cell wall into the cell where they are recognized by adequate receptors.
the changes in cuticular permeability , the ros production  and the diffusion of bioactive molecule through the surface  are all hallmarks of such a scenario.
the slightly increased permeability of myb96- <dig> shown in  <dig> permeability tests  seems to be sufficient to allow for faster ros production after inoculation and an increased resistance but not enough to allow sufficient leakage of active diffusates.
recent observations have shown a strong link between the presence of ros and resistance to b. cinerea after wounding, and the data presented here agree with these conclusions.
in fact, ros accumulation and resistance after wounding were shown to depend on calcium changes and all occur at the same location further supporting this hypothesis .
but the nature of the elicitors of these and corresponding receptors remain to be determined.
sensing of stretch  by mechano-sensitive proteins, for example by stretch-activated channels in the membrane might be another way sms is perceived and transduced.
a possible model is that mechanical stimulus at the surface of the cell stretches such channels initiating a calcium flux  <cit> .
the sms-induced transient burst of calcium  and the induction of genes  that were previously associated with the perception of mechanical stimuli  <cit>  would argue in favor of this scenario.
but sms like cutinase- or wound-induced resistance to b. cinerea are independent of ja signaling  <cit>  , an observation that would differentiate sms from a recent study on induced resistance to b. cinerea induced by leaf bending  <cit> .
leaf bending  also referred to as gentle touch is only accompanied by a ca 30% reduction in lesion after inoculation with b. cinerea and is ja-sensitive  <cit> .
this contrasts with the present results where sms was observed to lead to a full immunity to b. cinerea that is insensitive to ja.
experiments would now be needed with mutants blocked in mechano-sensitive touch receptors to differentiate between these pathways.
it is most likely that sms also leads to a major cellular reorganization such as that described by hardham and colleagues   <cit> ; the cellular details of the perception and attending mechanisms await now further studies.
CONCLUSIONS
wounding and sms exemplify how plants can react to a situation where in principle they might become more vulnerable.
they rely on the deployment of a stress response that includes rapid changes in calcium levels and the release of active molecules such as ros that subsequently lead to the activation of defense reactions exemplified by a strong resistance against the virulent b. cinerea.
interestingly, sms is not associated with wounding and a modification in the wax layer is enough to produce this syndrome.
the fact that sms is also leading to the induction of so-called touch genes leaves two possible scenarios open: i) modifications of the plant surface by sms lead to a facilitated perception of damps or mamps by membrane receptors with subsequent activation of defenses or ii) sms is perceived by mechano-sensors that subsequently initiate resistance .
overall, these results highlight the remarkable ability of plants to sense external mechanical stimuli and activate a powerful defense response.

 plants are exposed to various forms of mechanical stress caused by rain, snow, wind, animals, pathogens or plants themselves. such mechanical stimuli induce responses in the plant that were shown in many cases to have an adaptive value. a classical example is the response of trees to wind that results in shorter and thicker trunks. reaction or compression wood is an anatomical consequence of sensing mechanical stress with subsequent lignification of cell walls. plants also respond to a more delicate mechanical stress referred to as touch that leads to nastic or tropic responses. classical examples include the folding of mimosa pudica’s leaflets, the leaf closure of the venus fly trap or the coiling of tendrils. such stimuli lead to visible responses such as a reorientation of organs or changes in shape allowing catching an insect or improved anchorage. the response of plants to mechanical stimuli can also be more discrete without any apparent overt changes. for example, mechanical stress associated with damage or wounds can lead to increased resistance to insects or fungal pathogens. a closer look at the response to wounding has shown that it induces biochemical and molecular changes often associated with subsequently induced resistance mechanisms. for instance, wounded plants produce reactive oxygen species, undergo changes in lignification, ja, other hormones or wound signals and exhibit changes in gene expression that are associated with induced defense reactions. in a previous study we have shown that wounding of arabidopsis thaliana leaves induces a strong and transient immunity to botrytis cinerea the causal agent of grey mould. the expression of genes for camalexin biosynthesis and of glutathione-s-transferase, the activity of a map kinase activity and the accumulation of camalexin are primed by wounding. wound-induced immunity is independent of the major plant defense pathways involving salicylic acid, ja or ethylene, but depends on glutathione. recently, we have shown that wounding leads to the formation of ros within minutes and ros are required for wound–induced resistance to b. cinerea. a strong constitutive resistance to b. cinerea also takes place in mutants such as bdg and lacs defective in the production of a functional cuticle and displaying a phenotype of enhanced cuticular permeability. moreover, leaf surfaces treated with cutinase produced ros and became more protected to b. cinerea. thus, increased permeability of the cuticle is linked to ros formation and resistance to b. cinerea. in this study, we have further explored the responses of a. thaliana such as ros and resistance to b. cinerea in leaves that are subjected to more gentle form of mechanical stimulation.  sms treatment of a. thaliana leaves induces ros and resistance to b. cinerea we have treated leaves of a. thaliana by gently rubbing them between thumb and forefinger, a mechanical stress that is herewith referred to as a soft mechanical stimulation. the inoculation of leaves immediately after sms with spores of b. cinerea led to a strong decrease in lesion size. a rapid burst in ros evidenced by a green fluorescence was observed in leaves infiltrated with 5--carboxy- -dichlorodihydrofluorescein diacetate immediately after sms. dcf-da detects a broad range of oxidizing reagents including h2o and o2- and its use has been previously described. sms-induced resistance as well as wound-induced resistance were still detected in mutants of nadph oxidase d and f as well as in the double mutant meaning that others rboh proteins are implicated in the formation of ros. the response to sms showed a dose-dependence. applying only one event of sms already lead to a small but detectable decrease in lesion size as well as production of discrete patches of green dcf-da-fluorescence. we used stimulations throughout this study as this lead to the strongest effects. unstimulated upper leaves of plants stimulated on one lower leaf were not protected against b. cinerea showing the absence of a systemic effect. moreover, wearing latex gloves was equally effective for sms-induced resistance. the sms-induced resistance to b. cinerea was transient: when plants were inoculated 8 h after sms, about 50% of the resistance was lost and h after sms, plants were fully susceptible. sms was followed by a rapid change in intracellular calcium as detected using both yellow cameleon- or aequorin-expressing plants. the expression of so-called touch genes previously associated with mechanical stimulation such as tch tch cml and cml was also induced to 1 h after sms treatment. the effect of sms could still be observed in mutants of ja biosynthesis and signaling. the ethylene mutant ein2- also responds to sms. sms is not accompanied by cellular damage we next examined the occurrence of overt wounding after sms, since wounding was shown previously to strongly affect resistance to b. cinerea. macroscopic signs of wounding were not visible on sms-treated leaves. nevertheless we assessed the effect of sms on the cell surface by scanning electron microscopy of live leaf surfaces as well as vital staining using trypan blue. sms-treated epidermal cells looked perfectly turgid when viewed under the sem. the waxy surface of some cells appeared slightly affected but the cells themselves retained turgidity. some trichomes and cells at their base displayed damage. when sms-treated leaves were stained using trypan blue, a vital dye that marks the presence of dead cells, epidermal cells were essentially intact 1 h after treatment with the exception of isolated cell groups that stained in blue at the basis of trichomes, in agreement with observations made using the sem directly after sms. thus, sms did not lead to massive cellular damage compared to wounding with forceps that was observed previously. the question remains if the damaged cells at the basis of the trichomes might constitute a sufficiently strong wound stimulus to induce ros and resistance. this question was approached using the trichome-less glabrous gl mutant of arabidopsis. sem images of leaf surfaces of the gl mutant were compared to wt plants and in both plants cells retained turgidity after sms. no damaged cells were observed using the vital stain trypan blue in gl mutant after sms. in fact, the untreated gl mutant is as susceptible to b. cinerea as wt plants and after sms, gl displayed resistance to b. cinerea to the same extent as the wild type. thus, sms induced resistance independently of the presence of trichomes and sms-induced resistance to b. cinerea is not based on wounding of cells. alterations in cuticular permeability and sms an accumulation of ros was previously observed in a. thaliana plants displaying increased cuticular permeability such as mutants defective in cuticle biosynthesis or in the formation of abscisic acid. therefore we have determined if the sms-stimulated leaves underwent a change in cuticular permeability. the permeability of the cuticle of sms-stimulated leaves was assessed using various diagnostic tests. sms-treated leaves displayed an increase in cuticular permeability as collectively indicated by increased chlorophyll leakage, retention of toluidine blue as well as calcofluor white staining. sms is not accompanied by changes in aba levels wound-induced resistance to b. cinerea is lost when wounded plants are not maintained under a humid environment. this loss is caused by aba, the level of which increases under dry conditions. in accordance, mutants impaired in aba were fully resistant after wounding, both when plants were maintained under humid or dry environments. interestingly, increase in resistance and production in ros were observed whether plants were maintained at humid or dry conditions after sms. consequently, we also determined possible changes in the level of aba. no changes were observed between the levels of aba after sms in plants maintained under humid or dry conditions. this marks a clear difference between sms and wound-induced resistance. sms and the surface wax layer since sms likely perturbs the waxy surface of the plant without much damage to the underlying cells, we explored the importance of the wax layer on the leaf surface. we have used the myb96- mutant affected in the transcription factor myb involved in the biosynthesis enzymes condensing very-long-chain fatty acids involved in cuticular wax biosynthesis. untreated myb96- mutants displayed increased resistance to b. cinerea. the ros response of myb96- mutants to b. cinerea was much faster than in the wild type since green fluorescence was detected already 3 hours after inoculation with b. cinerea. the myb96- mutant also displayed a slight modification of permeability as indicated by the toluidine blue and calcofluor white test, but this modification was not detected with the chlorophyll leakage test. thus, myb96- altered in the wax layer displays a somewhat similar syndrome although less obvious that the wild type leaves after sms treatment. sms and leaf diffusates changes in the permeability of the cuticle were shown to be associated with the leakage of diffusates that prevent the development of b. cinerea in vitro and in vivo. we have tested if bioactive diffusates can be obtained from leaf surfaces of sms-treated wt or myb96- plants. sms applied to gl and myb96- mutants was similarly effective as on sms-treated wt plants. without sms, diffusates collected from the surfaces of both gl and myb96- plants were inactive similarly to those from wt plants. thus sms acted on leaf surfaces in a comparable way in plants or mutants displaying increased cuticular permeability. discussion wounding inflicted by clamping leaves with forceps or puncturing with a needle induces a strong immunity of a. thaliana to b. cinerea. in this study, we have explored the effect of softer forms of mechanical stimulation on the resistance of a. thaliana to b. cinerea. in particular, we have observed that a gentle mechanical stimulus applied to the surface of the leaf induced a transient and localized resistance to b. cinerea. plants are known to be equipped with a sensitive and discriminative sensory system for the detection of molecular patterns generated by pathogens. here we show that a gentle mechanical stress can also be perceived in a differentiated way and lead to specific plant responses that include resistance against a virulent necrotrophic fungus. how does sms compare to wounding? overall, the results on sms-induced resistance to b. cinerea overlap with wound-induced resistance of a. thaliana leaves. the results presented here further the published observations on wounding by showing that a soft mechanical friction of the surface layer without wounding the underlying cell is already enough to induce resistance. the absence of overt cellular breakage after sms is supported by the absence of change in the levels of aba after sms under dry or humid conditions and the observation of leaf surfaces of wt and glabrous mutants after sms. despite this, sms-treated plants as well as the waxless mutant display an increased permeability to toluidine blue or calcofluor white, indicating that the cuticular barrier is affected to a certain extent. the altered cuticular permeability might allow the diffusion of a bioactive molecule observed in sms-treated plants, similarly as in cuticle-defective mutants. both wounding and sms lead to a rapid and important release of ros. a number of reports have associated mechanical stress with an increased production of ros. for example, rubbing tomato plant internodes results in a rapid and lasting accumulation of h2o. ros are well known for their effect as intracellular signals and were shown to be involved in the activation of defenses in response to biotic and abiotic stress. sms-induced resistance to b. cinerea still takes place in atrbohd, atrbohf and atrbohd/f mutants of nadph oxidoreductase making it unlikely that these nadph oxidoreductases are involved. it cannot be excluded that some other arabidopsis nadph oxidoreductases are involved. this is similar to results obtained with these mutants after wounding. in addition, sms- like wound-induced resistance are both independent of ja signaling. how is sms perceived by the plant? the results presented here lend themselves to a similar interpretation as the studies on resistance to b. cinerea observed in a. thaliana after wounding or in plants with defective cuticles. sms might modify the plant surface making it more permeable thus allowing a better transfer of damps or mamps through the cell wall into the cell where they are recognized by adequate receptors. the changes in cuticular permeability, the ros production and the diffusion of bioactive molecule through the surface are all hallmarks of such a scenario. the slightly increased permeability of myb96- shown in permeability tests seems to be sufficient to allow for faster ros production after inoculation and an increased resistance but not enough to allow sufficient leakage of active diffusates. recent observations have shown a strong link between the presence of ros and resistance to b. cinerea after wounding, and the data presented here agree with these conclusions. in fact, ros accumulation and resistance after wounding were shown to depend on calcium changes and all occur at the same location further supporting this hypothesis. but the nature of the elicitors of these and corresponding receptors remain to be determined. sensing of stretch by mechano-sensitive proteins, for example by stretch-activated channels in the membrane might be another way sms is perceived and transduced. a possible model is that mechanical stimulus at the surface of the cell stretches such channels initiating a calcium flux. the sms-induced transient burst of calcium and the induction of genes that were previously associated with the perception of mechanical stimuli would argue in favor of this scenario. but sms like cutinase- or wound-induced resistance to b. cinerea are independent of ja signaling, an observation that would differentiate sms from a recent study on induced resistance to b. cinerea induced by leaf bending. leaf bending also referred to as gentle touch is only accompanied by a ca 30% reduction in lesion after inoculation with b. cinerea and is ja-sensitive. this contrasts with the present results where sms was observed to lead to a full immunity to b. cinerea that is insensitive to ja. experiments would now be needed with mutants blocked in mechano-sensitive touch receptors to differentiate between these pathways. it is most likely that sms also leads to a major cellular reorganization such as that described by hardham and colleagues; the cellular details of the perception and attending mechanisms await now further studies.  wounding and sms exemplify how plants can react to a situation where in principle they might become more vulnerable. they rely on the deployment of a stress response that includes rapid changes in calcium levels and the release of active molecules such as ros that subsequently lead to the activation of defense reactions exemplified by a strong resistance against the virulent b. cinerea. interestingly, sms is not associated with wounding and a modification in the wax layer is enough to produce this syndrome. the fact that sms is also leading to the induction of so-called touch genes leaves two possible scenarios open: i) modifications of the plant surface by sms lead to a facilitated perception of damps or mamps by membrane receptors with subsequent activation of defenses or ii) sms is perceived by mechano-sensors that subsequently initiate resistance. overall, these results highlight the remarkable ability of plants to sense external mechanical stimuli and activate a powerful defense response. 
after gentle mechanical sweeping of leaf surfaces, a strong resistance to b. cinerea was observed. in a previous study we have shown that wounding of arabidopsis thaliana leaves induces a strong and transient immunity to botrytis cinerea, the causal agent of grey mould. in this study, we have further explored ros and resistance to b. cinerea in leaves of a. thaliana exposed to a soft form of mechanical stimulation without overt tissue damage. arabidopsis can detect and convert gentle forms of mechanical stimulation into a strong activation of defense against the virulent fungus b. cinerea. this was preceded by a rapid change in calcium concentration and a release of ros, accompanied by changes in cuticle permeability, induction of the expression of genes typically associated with mechanical stress and release of biologically active diffusates from the surface. this reaction to soft mechanical stress was fully independent of jasmonate. in addition, leaves exposed soft mechanical stress released a biologically active product capable of inducing resistance to b. cinerea in wild type control leaves. reactive oxygen species are formed within minutes after wounding and are required for wound–induced resistance to b. cinerea.
 in a previous study we have shown that wounding of arabidopsis thaliana leaves induces a strong and transient immunity to botrytis cinerea, the causal agent of grey mould. reactive oxygen species are formed within minutes after wounding and are required for wound–induced resistance to b. cinerea.  in this study, we have further explored ros and resistance to b. cinerea in leaves of a. thaliana exposed to a soft form of mechanical stimulation without overt tissue damage. after gentle mechanical sweeping of leaf surfaces, a strong resistance to b. cinerea was observed. this was preceded by a rapid change in calcium concentration and a release of ros, accompanied by changes in cuticle permeability, induction of the expression of genes typically associated with mechanical stress and release of biologically active diffusates from the surface. this reaction to soft mechanical stress was fully independent of jasmonate. in addition, leaves exposed soft mechanical stress released a biologically active product capable of inducing resistance to b. cinerea in wild type control leaves.  arabidopsis can detect and convert gentle forms of mechanical stimulation into a strong activation of defense against the virulent fungus b. cinerea. 