Introduction

The success of young life stages of plants, such as tree saplings, may be critical to establishment, especially in harsh environmental conditions. Saplings tend to have distinct physiological and morphological differences compared to adult plants, such as increased photosynthetic capacity, carbon allocation, and unique xylem considerations (Day et al. 2001; Niinemets 2002). As a result, juvenile age classes are generally considered more sensitive to environmental stress than mature trees (Smith et al. 2003; Greenwood et al. 2008). These differences may enable young life stages to maximize carbon uptake, growth, and carbon storage, thus over-winter survival. Ultimately, seedling and young sapling physiological success can dictate species distribution, particularly across ecotonal gradients such as forest tree lines (Clark et al. 1999; Germino et al. 2002; Bader et al. 2007).

The spruce–fir [Picea rubens Sarg. and Abies fraseri (Pursh) Poir.] forests of the southern Appalachian Mountains are considered remnants of the most southern boreal forest that dominated the lower elevations of the southeastern United States during the late Pleistocene. These mountain-top communities occur today only at seven locations throughout southern Virginia, western North Carolina, and eastern Tennessee (Oosting and Billings 1951; Ramseur 1960; White 1984; Delcourt and Delcourt 1984), existing within a mosaic of northern hardwood forests and grass bald communities (Whittaker 1956; Mark 1958). In the recent past, these relic forests have been threatened by stress factors such as logging, acid rain, attacks from invasive insects {e.g., balsam wooly adelgid [Adelges piceae (Ratzeburg)]}, and alteration of the environment due to climate change (White 1984; Arthur and Hain 1986; Bruck and Robarge 1988; Busing et al. 1988; McLaughlin et al. 1990; White and Cogbill 1992].

Cloudiness, specifically cloud immersion, is one environmental factor hypothesized as a major contributor to the continued existence of these boreal, spruce–fir forest communities at such southern latitudes (Cogbill and White 1991). Moreover, the mean altitude of the cloud base of the southern Appalachians has been associated with the current distribution and persistence of spruce–fir forests on these mountaintops (Braun 1964). Scientists consider the effects of clouds of primary importance in understanding the impacts of climate change on ecosystems, yet they are poorly understood (IPCC 2007; Ruddiman 2008). Climate models suggest that, in general, clouds will become less frequent and cloud ceilings will rise in the southern Appalachians with warmer air temperatures (Pounds et al. 1999; Still et al. 1999; Foster 2001; Richardson et al. 2003; Brient and Bony 2012). However, these model predictions vary widely depending on the algorithms applied and geographic location. In direct contrast to these predictions, Richardson et al. (2003) found that the cloud ceiling had lowered slightly over the past 30 years in the southern Appalachians, but that this change lies within the natural variability of cloud heights at that time scale. Other studies have reported relationships between temperature, radiative forcing, and cloud patterns, but these results also vary by region (Croke et al. 1999; Gregory and Webb 2008). Undoubtedly, cloud patterns seem to remain one of the greatest sources of uncertainty in the IPCC predictions of climate change impacts (IPCC 2007).

Mountains that commonly experience clouds, including cloud immersion, occur on a broad geographic scale (Bruijnzeel et al. 2010) and have been found to generate less intense sunlight, a greater proportion of diffuse light, decreased air temperatures, and higher ambient humidity levels (Gu et al. 2002; Letts and Mulligan 2005; Min 2005). The current paper examines the difference in microclimate and the resulting plant physiology between two elevations with a specific focus on cloud immersion. Few studies have reported comparable measurements of microclimate during cloud immersion with an additional comparison of the variation between upper and lower elevation limits (Young and Smith 1983; Knapp and Smith 1990; Johnson and Smith 2008; Reinhardt and Smith 2008b). This is of additional importance in the southern Appalachian spruce–fir forests because their distribution is confined to only ~500 m in elevation between their upper (1,960 m) and lower (1,510 m) limits. The maximum elevation possible in the southern Appalachians is 2,037 m, the height of Mt Mitchell.

Photosynthesis, transpiration, leaf conductance, and water status (xylem water potentials) were measured for saplings of P. rubens and A. fraseri at their lowest elevational (LE; 1,510 m) and highest elevational (HE; 1,960 m) limits. Photosynthetic photon flux density (PFD), air temperature, and vapor pressure deficit (VPD) were also measured continuously at each site. We hypothesized that cloud-immersed days would result in greater leaf conductance and photosynthetic carbon gain, especially in the afternoon on clear days when stomata typically begin to close in response to greater water stress. Leaf transpiration should also be reduced on cloud-immersed days, causing greater plant water potentials, leaf conductance, and photosynthesis by midday and during the afternoon. We also expected that the LE sites would experience less frequent cloud immersion and, thus, higher temperatures and transpiration, resulting in greater reductions in leaf conductance and photosynthesis due to even more negative xylem water potentials than the HE sites. Because A. fraseri tends to dominate the community at higher elevations (Whittaker 1956), greater effects in this species at the LE sites were anticipated.

Materials and methods

To examine the difference in photosynthetic gas exchange and water status, field plots were established at the HE and LE limits of Southern Appalachian spruce–fir communities. Measurements were taken over a summer growth season (early, middle, and late) of 2011. The HE and LE sites were alternated between being measured first or second, and were both measured within 1 h of each other, starting at the specified time. All data were compared between the LE and HE plots, between clear and cloud-immersed days, and between species (Abies fraseri and Picea rubens).

Study sites

Study areas were located in the spruce–fir forest of the Black Mountains near Mt. Mitchell, North Carolina (Mt. Mitchell State Park, 35°45′53″N, 82°15′54″W), the highest point in the eastern United States (2,037 m). The Black Mountains are dominated by P. rubens (red spruce) and A. fraseri (Fraser fir) above ca. 1,500 m elevation and are characterized by high rainfall and snowfall (>1,800 mm year−1 evenly distributed through the year), moderate temperatures (13.5 °C mean maximum air temperature during the growth season, May up to and including September) and frequent cloud cover and immersion (Mohnen 1992; Reinhardt and Smith 2008a). All physiological measurements were taken on saplings of both study species along a transect at 1,510 m (35°71′N, 82°27′W) and 1,960 m (35°76′N, 82°26′W) elevation. Transects were selected on south-facing slopes in forest sites that were considered representative of the forest type at that elevation, i.e., a mostly closed canopy (>80 %) composed of both tree species and a ground layer of ferns and herbs, plus some dead trees due, most likely, to balsam wooly adelgid death (Pittillo and Smathers 1979; Bruck and Robarge 1988; Goelz et al. 1999). Healthy saplings between 0.7 to 1 m in height and without noticeable cone production were selected along transects that were considered representative of the mature spruce–fir communities, although some important differences between the LE and HE sites should be noted. At the LE site, spruce dominated both the sapling and canopy communities, canopy trees were generally taller (~12 versus ~8 m), and the canopy was slightly more closed [12 versus 18 % canopy openness determined using a hand-held spherical convex densitometer (model A; Forestry Suppliers, Jackson, MS)]. Both sites were likely harvested for timber before the development of Mt. Mitchell state park (1915) and Pisgah national forest (1916), making the current, second-growth forest around 100 years old.

Microclimate and cloud regime

Air temperature, humidity, and photosynthetic PFD were measured continuously throughout the entire growing season. At each site, three sensors measuring photosynthetic PFD (μmol m−2 s−1, 0.4–0.7 μm wavelengths) were placed at approximate sapling heights (1 m above ground) and spaced ca. 10 m apart. Measurements of PFD were logged from sunrise to sunset every 15 min from 23 May 2011 to 28 September 2011 using Photosynthetic Light Smart Sensors (model S-LIA-M003; Onset, Bourne, MA) connected to four-channel HOBO Micro Station data loggers (model H21-002; Onset). All PFD sensors were matched and calibrated against a recently calibrated (at factory) LI-COR quantum sensor (model 190S; LI-COR, Lincoln, NE), as well as a Science Associate precision solarimeter (model 1240), before installing in the field. Air temperature and relative humidity were measured every 15 min at 1 m above the ground using a HOBO Pro v2 sensors and data logger (model U23-001; Onset). Calibration of the temperature/humidity sensor located at the central location was checked periodically with well-ventilated and shielded, fine-wire (36 ASW gauge) thermocouple psychrometers under field conditions. The temperature/humidity sensor was placed in an open area for greater air flow and was shielded from direct sunlight through the entire day by a plastic shield painted with 3M white–velvet paint (total solar reflectance >90 %) and mounted ~5 cm above the sensors.

To verify cloud immersion for each measurement day, the study utilized an eastward-facing web camera mounted at the park office (North Carolina Division of Parks and Recreation) located at ~1,900 m elevation and pointed at the forest edge displaying everything above mid-canopy level. Because the camera was not located precisely at the field measurement sites, camera images were compared with PFD and humidity values to confirm estimates of canopy immersion for a given time period. Each morning and afternoon was classified as cloud immersed, cloudy, or sunny by requiring at least 2 h of each condition [see Berry and Smith (2012) for details of specific classification criteria]. While measurements were made on other day types as well, this study compares only days categorized as cloud immersed for both morning and afternoon time periods with those classified as sunny during the same periods.

Leaf gas exchange and water relations

Plant xylem pressure potentials (Ψ), photosynthesis, leaf conductance, and transpiration were measured three times (early, middle, and late) during the growing season of 2011. Measurements at both LE and HE sites for both species were taken on 2–3 completely cloud-immersed days and 2–3 completely clear days during May, July, and September 2011. Xylem pressure potential measurements were taken at 0600 hours (pre-dawn), 1000 hours (morning), and 1500 hours (afternoon) and included five replicates of shoots 4–6 cm in length and located on the south-facing sides of each individual sampled. Plant water status was determined by measuring Ψ for saplings of each species using a Scholander-type pressure chamber (model 1000; PMS Instrument, Corvallis, OR).

Leaf gas exchange was measured on the same individuals and days as Ψ measurements using a LI-COR LI-6400 model portable photosynthesis system (LI-COR). Measurements were made at 0900 hours (morning) and 1400 hours (afternoon) on previous-year shoots on south-facing sides of the sapling at mid-tree heights. During measurement, the natural orientation of shoots was maintained and air temperature and relative humidity inside the leaf chamber were regulated to be within ±5 % of ambient values. This was accomplished by measuring relative humidity outside the chamber and setting the relative humidity of the incoming air to this value. The instrument maintains a flow rate to obtain the desired humidity. When measurements were made during cloud immersion, needles were thoroughly blotted dry with tissue paper immediately before measurement (Brewer and Smith 1997). Leaf conductance values were monitored very closely and all gas exchange measurements were discarded if conductance seemed inaccurately high due to moisture still on the leaf surface. On cloud-immersed days, PFD was <400 μmol m−2 s−1, the leaf-to-air vapor pressure difference (LAVD) remained <0.7 kPa, and leaf temperatures were within 0.5 °C of ambient air temperature during morning and afternoon measurement periods. On sunny days, PFD ranged from 700 to 1,000 μmol m−2 s−1 during morning measurements and 1,200–1,700 μmol m−2 s−1 during afternoon measurements. The LAVD remained >1.2 kPa at HE and over 1.5 kPa at LE, while leaf temperatures were within 1 °C of ambient air temperature [see Reinhardt and Smith (2008a) and Berry and Smith (2012) for further details and data]. The LI-COR LI 6400 clear conifer chamber was used for all measurements with total leaf area in the cuvette calculated by counting the number of leaves in the chamber and using a mean total leaf area determined as described in Smith et al. (1991).

Statistics

All data comparisons were tested for statistical significance using ANOVA according to the appropriate tests for normality and equality of variance (Zar 1999). Specifically, ANOVA was used to compare differences between day type, elevation, species, time of day, and month. When the assumptions (normality and/or equality of variance) of ANOVA were not satisfied, non-parametric tests were employed using Wilcoxon rank sum tests with the alpha value recomputed using the Bonferroni correction. Throughout the text, when ANOVA was used an F-value and probability are presented, and when the Wilcoxon rank sum test was used a χ2-value and probability are presented. For all gas exchange measurements, there was no significant effect of the time of year that measurements were taken, and thus all months were grouped for statistical analysis. All analyses were conducted using JMP (SAS, Cary, NC) and figures were constructed using SigmaPlot v. 11 (Systat Software, San Jose, CA).

Results

Microclimate

The HE sites had more frequent cloud immersion with 75 % of days having at least 30 min of cloud immersion (56 % for LE). As expected, the LE sites had higher maximum air temperatures (18.4 versus 15.5 °C; F 1,190 = 85.4, P < 0.0001), VPDs (1.78 versus 1.52 kPa; F 1,190 = 39.9, P < 0.0001), and lower cumulative PFD (3.5 mol m−2 day−1 versus 10.4 mol m−2 day−1; F 1,190 = 110.2, P < 0.0001) than the HE sites. Cloud-immersed days at the HE sites were significantly cooler with a mean daily temperature of 15.2 °C compared to 16.6 °C on clear days (Table 1; F 1,94 = 8.75, P = 0.004). Cloud immersion resulted in lower mean cumulative daily PFD (7.3 mol m−2 day−1 versus 10.4 mol m−2 day−1; F 1,94 = 25.1, P < 0.0001) and mean daily VPD (1.40 versus 1.57 kPa; F 1,94 = 5.7, P = 0.019) (Table 1). The VPD values were higher than expected during cloud-immersed days because their mean values incorporate some data points during the day when complete cloud immersion was temporarily absent. At the LE sites cloud immersion also resulted in a lower mean temperature (17.8 versus 19.1 °C; F 1,94 = 9.12, P = 0.003), lower cumulative daily PFD (2.9 mol m−2 day−1 versus 4.5 mol m−2 day−1; F 1,50 = 27.6, P < 0.0001), and lower VPD (1.66 versus 1.87 kPa; F 1,94 = 15.3, P = 0.0002). Days with different cloud regimes were evenly distributed across the entire study period.

Table 1 Mean daily air temperature, vapor pressure deficit (VPD), and cumulative daily photon flux density (PFD) on cloud-immersed and clear days at the upper (1,960 m) and lower (1,510 m) elevational extent of Southern Appalachian spruce–fir communities during the summer growing season
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Photosynthetic gas exchange

There was a significant effect of cloud immersion on leaf-level photosynthesis in both species and elevations (Fig. 1; Table 2; χ2 = 5.93, P = 0.015). Afternoon photosynthesis was significantly greater on cloud-immersed days than clear days at both elevations (HE, χ2 = 22.29, P < 0.0001; LE, χ2 = 22.75, P < 0.0001). For example, in May, HE A. fraseri had an afternoon photosynthesis of 0.63 μmol m−2 s−1 on clear days and 1.96 μmol m−2 s−1 on cloud-immersed days (Fig. 1a, b). Similarly for P. rubens in May, mean afternoon photosynthesis on cloud-immersed days was 1.00 μmol m−2 s−1 versus 0.78 μmol m−2 s−1 on clear days (Fig. 1b). Normalizing photosynthesis relative to PFD (photosynthesis/PFD) gave values greater during cloud-immersed periods. On clear days, morning ratios ranged from 0.0022 to 0.0031 and afternoon ratios ranged from 0.0003 to 0.0005, which were considerably lower than morning values. On cloud-immersed days these values were more similar between morning and afternoon (0.0034–0.0060 and 0.0027–0.0050, respectively). Photosynthesis at LE was always lower than at HE, but followed the same trend (χ2 = 65.89, P < 0.0001). For example, LE A. fraseri had a mean afternoon photosynthesis in May of 0.42 μmol m−2 s−1 on clear days versus 0.67 μmol m−2 s−1 on cloud-immersed days (Fig. 1). Morning (0900 hours) measurements tended to be similar across day types within each elevation and species, and no significant differences occurred between monthly measurements (χ2 = 5.06, P = 0.08).

Table 2 Percent change from morning to afternoon values of photosynthesis (A), transpiration (E), leaf conductance (g), plant water potentials (Ψ), and instantaneous water use efficiency (WUE = A/E) for Abies fraseri and Picea rubens
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Fig. 1
figure 1

a–f Net photosynthesis (A net) during morning (0900 hours) and afternoon (1400 hours) on saplings of Abies fraseri (triangles) and Picea rubens (circles). Measurements were taken simultaneously near the top (1,960 m) and bottom (1,510 m) of the communities’ elevational extent on five individuals each day and repeated for 2–3 days of each day type. Solid symbols and lines represent high elevation measurements and open circles with dashed lines represent low elevation measurements. Bars represent SE

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Leaf conductance followed a similar trend with afternoon measurements being significantly greater on cloud-immersed days (Fig. 2; Table 2; HE, χ2 = 29.25, P < 0.0001; LE, χ2 = 30.21, P < 0.0001). On clear days, afternoon values were under 0.1 mol m−2 s−1 for both species at both elevations. On cloud-immersed days, afternoon conductance varied between 0.20 and 0.35 mol m−2 s−1 at HE and between 0.13 and 0.20 mol m−2 s−1 at LE (Fig. 2). As expected, afternoon conductance was significantly lower than morning values (χ2 = 123.08, P < 0.0001) and, as with photosynthesis, morning measurements tended to be similar between clear and cloud-immersed days without significant differences between months (χ2 = 3.63, P = 0.16).

Fig. 2
figure 2

a–f Leaf conductance measurements during morning (0900 hours) and afternoon (1400 hours) on saplings of A. fraseri (triangles) and P. rubens (circles). Measurements were taken simultaneously near the top (1,960 m) and bottom (1,510 m) of the communities’ elevational extent on five individuals each day and repeated for 2–3 days of each day type. Solid symbols and lines represent high elevation measurements and open circles with dashed lines represent low elevation measurements. Bars represent SE

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Transpiration and plant water status

There was also a significant, negative association between cloud immersion and transpiration (Fig. 3; Table 2; χ2 = 113.32, P < 0.0001). In contrast to leaf conductance and photosynthesis, differences in transpiration were much more pronounced for morning (0900 hours) measurements, i.e., less than 0.3 mmol m−2 s−1 on cloud-immersed days compared to 1.2 and 1.5 mmol m−2 s−1 on clear days (χ2 = 44.29, P < 0.0001) at both the LE and HE sites. Differences in afternoon transpiration were less pronounced, but still significant (Fig. 3; HE, χ2 = 32.85, P < 0.0001; LE, χ2 = 7.36, P = 0.007). A decrease from morning to afternoon measurements was less severe on cloud-immersed days (Table 2). Overall, cloud-immersed transpiration was similar to clear-day transpiration only when leaf conductance was low.

Fig. 3
figure 3

a–f Transpiration measurements during morning (0900 hours) and afternoon (1400 hours) on saplings of A. fraseri (triangles) and P. rubens (circles). Measurements were taken simultaneously near the top (1,960 m) and bottom (1,510 m) of the communities’ elevational extent on five individuals each day and repeated for 2–3 days of each day type. Solid symbols and lines represent high elevation measurements and open circles with dashed lines represent low elevation measurements. Bars represent SE

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On cloud-immersed days, xylem water potential (Ψ) improved from morning to afternoon measurements, whereas on clear days it declined continuously into the afternoon (Fig. 4; Table 2). Also, this increase in Ψ on cloud-immersed days was more pronounced at LE sites than HE sites. For example, A. fraseri Ψ in May at HE sites improved by 0.11 MPa, while no change occurred at LE sites. There was a significantly lower Ψ (χ2 = 77.58, P < 0.0001) at the LE sites (Fig. 4). At both elevations, predawn Ψ values were statistically similar between clear and cloud-immersed days (HE, χ2 = 0.04, P = 0.84; LE, χ2 = 1.65, P = 0.20).

Fig. 4
figure 4

a–f Xylem pressure potentials (Ψ) during predawn (0600 hours), morning (1000 hours), and afternoon (1500 hours) measurements on saplings of A. fraseri (triangles) and P. rubens (circles). Measurements were taken near the top (1,960 m) and bottom (1,510 m) of the communities’ elevational extent on five individuals each day and repeated for 2–3 days of each day type. Solid symbols and lines represent high elevation measurements and open circles with dashed lines represent low elevation measurements. Bars represent SE

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Differences in Ψ between clear and cloud-immersed days were most pronounced during the afternoon in both species (Fig. 4; χ2 = 4.34, P = 0037). For example, afternoon Ψ was 0.30–0.55 MPa greater on cloud-immersed days than clear days at high elevation, and was less pronounced at low elevation (0.05 and 0.25 MPa greater). Although there was a significant decrease in Ψ from May to September in all measurements (χ2 = 228.71, P < 0.0001), all differences between clear and cloud-immersed days were statistically similar across months.

Discussion

Microclimate

Our goal was to examine the effect of cloud immersion on photosynthetic gas exchange and water status in P. rubens and A. fraseri saplings at their lower and upper elevational limits of occurrence. The HE plots had more frequent cloud immersion (75 versus 56 %) and corresponding changes in air temperature, VPD, and PFD (Table 1). Cloud immersion was associated with a ~1.5 °C decline in mean daily air temperature at both elevational plots. Similarly, mean daily VPD was reduced by ~0.2 kPa and total daily PFD was reduced by 1.5–3.0 mol m−2 day−1 on cloud-immersed days (Table 1). However, LE cloud-immersed days still had higher VPD (1.66 versus 1.57 kPa) and temperature (17.8 versus 16.6 °C) than HE clear days.

Leaf conductance and photosynthesis

Afternoon leaf conductance and photosynthesis values were greater on days with prolonged cloud immersion than clear days, ranging from 30 to 230 % (Figs. 1, 2; Table 2). Stomata remained open well into the afternoon on cloud-immersed days, along with higher photosynthetic carbon gain. Conductance and photosynthesis on cloud-immersed days were also greater at the HE than LE sites (Figs. 1, 2). The down-regulation of afternoon photosynthesis on sunny days is likely due to the higher transpirational demands earlier in the day leading to lower xylem water potentials and partial stomatal closure (Urban et al. 2012).

In addition to enhanced stomatal opening, another possible explanation for higher photosynthesis in understory saplings, is the alteration in the directional nature of the incident sunlight. Cloud immersion increases the diffuse component of direct, incident sunlight which generates deeper penetration into the forest canopy and can increase canopy photosynthesis per unit ground area (Campbell and Norman 1998; Min 2005; Urban et al. 2007; Dengel and Grace 2010). Once diffuse light reaches the understory, Ishii et al. (2012) found that needle arrangement in Picea species utilized diffuse light effectively at a wide range of incoming angles. Also, diffuse light contains a higher portion of blue wavelengths than direct-beam sunlight, potentially stimulating greater stomatal opening, as found in Picea species at relatively low PFD levels (0–300 μmol m−2 s−1; Morison and Jarvis 1983). Measured light response curves for photosynthesis showed a light saturation point of approximately 200–400 μmol m−2 s−1 in both A. fraseri and P. rubens, supporting the idea that both study species can maximize photosynthesis in low levels of diffuse light such as found during cloud immersion (Reinhardt and Smith 2008a). While low light saturation values often lead to lower light levels for initiating photoinhibition, photosynthetic light response curves for these two species showed no evidence of photoinhibition up to 2,000 μmol m−2 s−1 (Johnson and Smith 2006), well above the maximum PFD in this study (1,700 μmol m−2 s−1). Also, a quantitative evaluation of changes in photosynthesis versus leaf conductance found a strong linear relationship (clear sky r 2 = 0.83, cloud immersed r 2 = 0.72) suggesting that the afternoon decline in photosynthesis on clear days was tightly coupled to corresponding decreases in leaf conductance (data not shown).

The results above are similar to several other studies examining the effect of cloud and light quality on plant ecophysiology. Results reported by Urban et al. (2007) and Dengel and Grace (2010) showed enhancement of leaf conductance in Abies and Picea species under cloudy and overcast conditions, but did not measure complete cloud immersion. Simonin et al. (2009) did find an increase in photosynthesis in Sequoia sempervirens as a result of fog. Tropical montane cloud forest studies have suggested that lower PFD from clouds would result in reduced photosynthesis (Bruijnzeel and Veneklaas 1998; Letts and Mulligan 2005). These differences could perhaps be due to the differential light requirements of conifers and tropical broadleaves.

A potential limitation of cloud immersion to gas exchange would be the prolonged formation of water films on leaf surfaces. Smith and McClean (1989), Ishibashi and Terashima (1995), and Letts and Mulligan (2005) reported large decreases in photosynthesis as a result of moisture and water-film formation on leaf surfaces, presumably because of the much lower diffusivity of carbon dioxide in water versus air (ca. 104 times slower in air). While there clearly was moisture on needles of both species during cloud immersion, a hydrophobic epicuticular wax layer, a small and curved needle shape, and a steep leaf inclination results in water beads that are easily shed (relatively low retention; Reed and Smith 2012) preventing any water film formation.

Transpiration and plant water status

During cloud immersion, transpiration remained below 0.5 mmol m−2 s−1 in both morning and afternoon measurements. This value was substantially lower than on mornings of clear days (1.5 and 2.0 mmol m−2 s−1) and even lower compared to afternoon transpiration values (Fig. 3; Table 2). The low VPDs during cloud immersion were more tightly coupled with transpiration rate than leaf conductance because conductance was high during both the morning and afternoon measurements. Continued low transpiration throughout the morning hours is a likely factor driving the high afternoon conductance values in response to greater Ψ. These data are corroborated by several other studies that reported a reduction in transpiration during cloud-immersed conditions (Graham et. al 2003; Burgess and Dawson 2004; Johnson and Smith 2008).

As mentioned above, cloud immersion probably led to improved plant Ψ, particularly by afternoon. On cloud-immersed days, there was an afternoon rebound in Ψ at HE sites by an average of 0.13 MPa, and as much as 0.25 MPa from morning to afternoon measurements (Fig. 4; Table 2). While this improvement seems minimal, it becomes more meaningful compared to clear days when a continued decrease in plant Ψ by a mean 0.14 MPa resulted in a mean decline of 0.27 MPa in the afternoon between clear and cloud-immersed days. Although information on this is growing, few studies have reported a recovery in plant Ψ over a daytime or the association with cloud immersion (Brodribb and Holbrook 2004; Johnson et al. 2009; Berry and Smith 2012).

Instantaneous water use efficiency (A/E) from morning to afternoon measurements actually showed decreases in WUE on cloud-immersed days and increases on clear days (Table 2), similar to the results of Reinhardt and Smith (2008a). During all measurements C i/C a values averaged 0.81 ± 0.06, within the standard range previously reported near our sites (Reinhardt and Smith 2008a). The change in WUE was likely the result of a greater decrease in photosynthesis than transpiration from morning to afternoon on cloud-immersed compared to clear days, although both values were reduced (also see Reinhardt and Smith 2008a).

Effect of elevation

Leaf photosynthesis, conductance, transpiration, and plant Ψ in both species were greater at HE sites compared to the same species at LE sites, especially for the endemic A. fraseri. For example, afternoon photosynthesis on cloud-immersed days averaged 0.9 μmol m−2 s−1 at HE sites compared to only 0.4 μmol m−2 s−1 at LE sites (Fig. 1). Additionally, there were smaller changes from morning to afternoon measurements in photosynthesis, transpiration, and Ψ at HE sites particularly on cloud-immersed days (Table 2). These increased values at high elevation suggest that both species, but particularly A. fraseri, are better suited to the microclimate at HE sites where more frequent cloud immersion occurred. This difference in microclimate between the two elevations includes an increase in the number of cloud-immersed days per growing season and a significant reduction in mean daily temperature and VPD (Table 1).

Conclusion

To our knowledge, data comparing gas exchange traits between the upper and lower elevational limits of P. rubens and A. fraseri and for clear and cloud-immersed days did not previously exist. The present study showed increases in photosynthesis, conductance, and xylem water potentials during cloud immersion, particularly in afternoon measurements at HE sites. Cloud-immersed days resulted in higher leaf conductance in both species during the afternoon measurement period, corresponding to increased photosynthesis with a stronger response at HE sites. Cloud immersion also corresponded to substantially reduced transpiration throughout the entire day leading to an improved Ψ. Xylem water potentials showed a significant improvement from morning to afternoon measurements demonstrating a role for cloud immersion in lessening daily plant water stress. Moreover, LE sites had reduced conductance and photosynthesis, increased transpiration, and more negative Ψ than the plots at the high elevation boundary of these species. Overall, both species experienced improved carbon gain and plant water status on cloud-immersed days. Also, the endemic Abies fraseri showed the greatest improvement in gas exchange physiology as a result of cloud immersion and seems particularly vulnerable at LE sites.

Understanding the ecophysiology of saplings may be critical in determining the limitations to regeneration in these already threatened forests, as well as potential shifts in elevational occurrence. Climate models predict an increase in summer temperatures of at least 3 °C (and up to 6 °C) by 2100 in moderate scenarios (IPCC 2007). These temperatures would result in southern Appalachian mountain peaks having air temperatures comparable to locations where spruce–fir communities currently do not exist (Pounds et al. 1999; Still et al. 1999; Foster 2001; Richardson et al. 2003; Brient and Bony 2012). If clouds become less frequent and cloud ceilings rise, as predicted by some climate models, the existence of southern Appalachian spruce–fir communities could be severely threatened. Because spruce–fir communities seem to be reliant on these cloud-immersed days for improved carbon gain and water conservation, it is likely that they will shift upward in elevation if cloud ceilings rise. The ultimate survival of this relic, refugial forest could certainly depend on the magnitude of future changes predicted for cloud regimes in these mountains.