From my experience with tobacco (including drought stress experiments) I would assume that your plants are not water stressed (if values represent midday data). In previous studies I found RWC values of around 75% at midday for well watered tobacco plants, irrespectively of growing indoors or outdoors (e.g. Galle et al. JXB 2009).

Relative water content of 75%  definitly suggest a  water  stress, at least  a mild one.

Based on the leaf water relationss, particularly RWC of 75%, the plants would have been drought stressed. I would say they were moderately stressed.

Well watered plants have normally 0 /-0,5 MPa. Your RWC result indicate drought stress, but the question is that normally or not for tobacco plants? Stress tresholds are changing between plant species.

A RWC of 74-77% would typically indicate shortage of water, but it is not the absolute figures that are important, rather the difference in water status parameters between your "drought" treatment and your "control" plants. For your experiment you can say your plants are under "drought" stress only if the parameters, such as RWC are significantly lower (more stressed) than for your control plants.

It will depend how "well-watered" your control plants are. If they are not at full hydration (RWC close to 100%), then maybe a RWC of 74-77% will not be significantly different from the level of stress in your controls. Once you know that your "control" and "droughted" plants differ significantly in some aspect of plant water status, then you can start to look for the consequences of those differences, at whatever scale you are working at - gene transcription to morphology.

Hope that helps. Steve

Although the rwc  indicates some stress, the cell pressure indicates the inverse. And these pressure levels are somewhat high, indicating plenty water.

For water potential, -1,5MPa is the "bottom" reference value for cultivated plants, but herbs vary between -1,0 to-2,5, or something near this. I really don't know the reference value for tobacco.

Dear J.D. Franco-Navarro

1) To my knowledge the your RWC indicate some dehydration status, but I do not know the sensitivity degree of your genotypes to dehydration. 75% could be quite a stress or not so much depending as well on the genotypes

2) Furthermore your water potential (predawn or midday?) is not very negative; in my opinion could indicate, at most, a moderate stress. Also, if this water potential is related to predawn values in fact your plants are under stress (and will reach even more negative values along the day time). If such water potential value is related to midday, the stress would seem to be only moderate. In that case the predawn would be in a range possibly (I just can guess) -0.5-1.0 MPa, which for vegetables (at least not for trees and schrubs) would configure a mild stress.

3) Visual observation (and touch of the leaves) constitute a quite good way of "analysis" of the wilting degree (and, thus, of the dehydration/stress to which the plant is exposed).

4) Finally, the stress intensity (and therefore the metabolic impact) it will depend as well on the duration (hours, days, weeks) of such dehydration value.

I hope that I could help you.

Best regards.

According to my experience, your plants are very well watered. A plant under water deficit can reach a RWC well below 50 %.

All of your parameters indicate mild stress, if you are interested to induce a strong drought stress into your plants I suggest to decrease drastically the watering for three-, four-, and six-week drought treatment. And add the parameters of dry weight into your determinations, as well the water content in your substrate to record the water depletion in your experiments.

Regards.

the level of RWC indicates that plants were quite stressed. But depends on the method of RWC measurement. If you used leaf discs, it is possible to estimate low RWC even for well-watered plants. Better to use whole leaf with petiole and dip only petiole to rehydrate the leaf. This way will give you better estimation of RWC than using leaf discs.

While I do agree with most of the above comments, I just want to draw your attention to pressure-volume characterization of tobacco by Nardini et al (Plant, Cell and Environment (2008) 31, 804–812 doi: 10.1111/j.1365-3040.2008.01798.x). Although it seems your plants were more acclimated and had lower osmotic potentials, your RWC measurement indicate moderate stress.

I also agree with others and with regard to the RWC plants are exposed to stress.

Tolerances to salt and water deficit were analyzed in transgenic tobacco plants that overexpress a UCP (AtUCP1) from Arabidopsis thaliana. Seeds of AtUCP1 transgenic lines germinated faster, and adult plants showed better responses to drought and salt stress than wild-type (WT) plants. These phenotypes correlated with increased water retention and higher gas exchange parameters in transgenic plants that overexpress AtUCP1. WT plants exhibited increased respiration under stress, while transgenic plants were only slightly affected. Furthermore, the transgenic plants showed reduced accumulation of hydrogen peroxide in stressed leaves compared with WT plants.

Furthermore,  your plants were more acclimated and had lower osmotic potentials,  its RWC measurement indicate that these plants subjected to moderate stress.

From my experience with tobacco (including drought stress experiments) I would assume that your plants are not water stressed (if values represent midday data). In previous studies I found RWC values of around 75% at midday for well watered tobacco plants, irrespectively of growing indoors or outdoors (e.g. Galle et al. JXB 2009).

Wow thank you all, I didn't expect any kind of answer, but that is awesome!!

Tobacco is a complex plant. I subjected my plants to well-watered irrigation treatment, keeping plants to 100% field capacity all the time in semi-hydroponic conditions, that's why I can't understand a stress. Maybe evapotranspiration was higher than the uptake of water at midday?, but Pressure probes values (Zimmerman et al 2008) and quantum yield of the PSII, are indicating me that my plants are healthy. Plants didn't show wilting, I am working with drought stress too  in those tobacco plants (another experment different to this control experiment) and wilting syntomps appear under 60% field capacity.

I measured RWC and potential parameters at pre-dawn conditions, and the protocol was with leaf disc, as described in Barrs & Weatherley 1962. Potential parameters were recorded by using a dew-point microvolt-meter (model HR-33T, Wescor, UT, USA) and the C-52 sample chamber.

Thank you again!!!!!

I would like to thank you for your questions. First, drought is a complex trait. It has different divination. And the conditions play important role in plant situation if not good controlled. 

As, I did my Ph D on drought tolerance in wheat is was very difficult to control the water and quantity for that reason we have used a chemical methods to screen our plants such as KI spray.

No, your results of RWC indicate that your plants has  a relatively low stress.

Eventually I solved all the problems concerning this post (or RG question), and I was successful to publish in "Journal of Experimental Botany." You can access the open-access publication here: http://jxb.oxfordjournals.org/content/early/2015/11/23/jxb.erv502.full

I am very grateful for all for your comments. Sincerely.

JD