Global atmospheric C dioxide concentration ( [ CO2 ] ) has increased since the industrial revolution ( IPCC 2001, IPCC 2007 ) . Atmospheric [ CO2 ] in recent yesteryear was about 350 µmol mol-1 ( IPCC 2001, IPCC 2007 ) and the current sum may even be higher ( Tans 2010 ) . By the twelvemonth 2100, it is predicted that atmospheric [ CO2 ] would be between 730 and 1020 µmol mol-1 ( IPCC 2007 ) . Increased [ CO2 ] can impact works ecosystem ( Ceulemans and Mousseau 1994, K & A ; ouml ; rner 2006 ) , species profusion and composing ( Wang 2007, Langley and Megonigal 2010 ) . At the single works degree, increased [ CO2 ] by and large enhance leaf degree photosynthesis, lower stomatous conductance to H2O vapor ( gs ) and transpiration rate ( E ) , increase H2O and food usage efficiencies ( Dang and Cheng 2004, Ainsworth and Rogers 2007, Cao et Al. 2007, Huang et Al. 2007, Ambebe et Al. 2009 ) at least in the short term. The effects of increased [ CO2 ] on works growing and physiological procedures may nevertheless, be limited by other factors such as dirt temperature and alimentary position of the dirt ( Lukac et al. 2010 ) .
Increased in atmospheric [ CO2 ] is expected to be accompanied by a 1.4-5.8 oC addition in planetary air temperatures ( IPCC 2001 ) . Any addition in air temperature may ensue in increased dirt temperature ( Domisch et al. 2002 ) because both are determined by the energy balance at the land surface and positively good correlatives ( Zheng et al. 1993 ) . Soil temperature can impact works ‘s physiological procedures such as C arrested development, stomatous conductance, transpiration rate, alimentary consumption and alimentary re-translocation every bit good as C dioxide consumption ( Cai and Dang 2002, Dang and Song 2004, Pregitzer and King 2005 ) . The effects of increased atmospheric [ CO2 ] and increased temperatures are expected to be highest in the boreal wood ( IPCC 2007 ) , where works growing is limited by dirt temperature and alimentary supply ( Jarvis and Linder 2000 ) .
In the boreal forest, the three most of import factors that limit tree growing are soil temperature and the trees ‘ ability to gaining control of CO2 and foods ( Jarvis and Linder 2000 ) . At present climatic conditions, the boreal wood is characterised by low dirt temperatures ( Domisch et al. 2002 ) . However, there is grounds that the part has been warming up faster than other parts on Earth ( Serreze et al. 2000 ) . Increased atmospheric [ CO2 ] coupled with warmer temperatures may alter species composing, competitory ability and resource usage ( Stewart et al. 1998 ) . Two congenerous conifer species in the part that may be impacted greatly by these predicted alterations are black spruce ( Picea mariana ( Mill ) . B.S.P ) and white spruce ( Picea glauca ( Moench ) Voss ) . These two species are the most widely distributed conifers in the boreal wood of North America ( Nienstaedt and Zasada 1990, Sims et Al. 1990, Viereck and Johnston 1990, Haavisto and Jeglum 1995 ) . The Arctic Climate Impact Assessment ( ACIA ) clime theoretical accounts suggest that rapid heating of the boreal wood may non let the growing of commercially valuable white spruce while the spread of black spruce in western Canada ‘s boreal wood may be reduced on hapless sites ( Juday et al. 2005 ) . On the other manus nevertheless, the increased in dirt temperature and [ CO2 ] can widen the species turning season and increase their photosynthetic rate and biomass production every bit long as there is adequate foods and wet ( Str & A ; ouml ; mgren and Linder 2002 ) .
Black spruce grows on sites runing from dry littorals, crushed rocks and shoal dirts on bedrock through deep food rich mineral dirts on highlands to waterlogged alimentary deficient peatlands ( Haavisto and Jeglum 1995 ) . In Ontario ‘s boreal wood, 54 % of black spruce stands is on highland mineral dirts with 46 % on peatlands ( Haavisto and Jeglum 1995 ) . On the other manus, white spruce grows best on moist highland sites and on good aerated and good drained flood plains ( Nienstaedt and Zasada 1990 ) . On moist highland mineral dirts, the species co-occur with or without other species such as doodly-squat pine ( Pinus banksiana Lamb. ) ( Nienstaedt and Zasada 1990, Sims et Al. 1990, Haavisto and Jeglum 1995 ) . On peatlands and flood plains nevertheless, black spruce grows in association with white spruce merely if aeration is equal ( Nienstaedt and Zasada 1990, Viereck and Johnston 1990, Haavisto and Jeglum 1995 ) . The broad natural scope of the two species makes them ideal to analyze in the context of increased [ CO2 ] as temperature and alimentary position of the dirts are besides varied within the species range. Root growing of both species increase quickly at dirt temperatures above 10 oC ( Grossnickle 2000 ) . Soil temperature for optimal growing of black and white spruce seedlings under field conditions is between 19 and 21 oC ( Heninger and White 1974, Tyron and Chapin 1983, Grossnickle and Blake 1985, Odlum and Ng 1995 ) . Under controlled conditions, dirt temperature of 15 oC may be optimal both for species root growing ( Peng and Dang 2003 ) . A dirt temperature of 22 ( black spruce ) and 21 oC ( white spruce ) has besides been reported as the optimum for upper limit the species net photosynthetic rate ( Dang and Cheng 2004 ) . Maximal alimentary consumption in spruce occurs at dirt temperatures of 20 oC under current CO2 degrees ( Gessler et al. 1998 ) . Soil temperatures above this cause diminution in root consumption of nitrate and ammonium ( Gessler et al. 1998 ) .
In this paper, we report on the physiological responses of boreal black and white spruce seedlings to the synergistic effects of dirt temperature and alimentary supply under current and projected atmospheric [ CO2 ] . The specific aims were to analyze the: ( I ) photosynthetic rate of both species under glandular fever and assorted growing conditions ( two ) in vivo activities of Rubisco under glandular fever and assorted growing conditions and ( three ) photosynthetic food usage efficiency of the two species at glandular fever and assorted growing conditions. Numerous surveies ( Grossnickle and Blake 1985, Grossnicke 2000, Cai and Dang 2002, Domisch et Al. 2002, Dang and Cheng 2004, Ambebe et Al. 2009, Cheng 2009 ) have reported on the effects of low dirt temperature on boreal tree seedlings physiology. A concern that remains to be answered is whether growing at elevated [ CO2 ] would do an upward displacement in the species current optimal dirt temperature in response to increase warming associated with higher [ CO2 ] . We attempt to turn to this by turning the seedlings in their current optimal dirt temperature and at a jutting heater temperature. It was expected that the species current optimal dirt temperature would switch upwards in response to increased [ CO2 ] and consequence in greater photosynthetic rate in both species, particularly when foods are equal.
If any addition in boreal forest productiveness in response to warmer temperatures and increased [ CO2 ] is to be maintained, it will hold to be accompanied by an addition in dirt food ( Lukac et al. 2010 ) . Surveies on works nutrition largely focus on N ( N ) because it is the food required in the largest measure and most likely to restrict C addition ( Chapin et al. 1987 ) . However, lacks of other alimentary elements have been reported ( Reich et al. 2009 ) . The rate of photosynthesis is dependent on leaf P concentration ( [ P ] ) ( Herold 1980 ) and its lack inhibits photosynthesis ( Lambers et Al. 2008, Reich et al. 2009 ) . It has besides been shown that P supply influences breakdown of N, including to Rubisco ( Warren and Adams 2002, Warren et al. 2005 ) . Leaf K ( [ K ] ) affects photosynthesis through ordinance of stomatous aperture ( Pallardy 2007 ) and any instability may ensue in reduced photosynthetic rate ( Pallardy 2007 ) . Specifically, P and K ( K ) lacks have been observed in black spruce ( Wells 1994, Teng et Al. 2003 ) and in white spruce ( Truong and Gagnon 1975 ) . To turn to such restriction, we maintained a changeless ratio of the three major alimentary elements ( N, P and K ) . Low alimentary supply normally limit works ‘s photosynthetic rate ( Aerts and Chapin 1999, Ainsworth and Long 2005 ) . However, increased [ CO2 ] can partly counterbalance for low alimentary supply in the short term, by diminishing the sum of Rubisco required to repair a given sum of C ( Percy and Bjorkman 1983 ) . While we expected that photosynthetic rate would be higher at elevated [ CO2 ] and high food supply, it was besides hypothesised that the negative effects of low alimentary supply on photosynthesis would be partially compensated for by elevated [ CO2 ] , particularly at the heater dirt temperature.
The stimulation of tree growing and map by enhanced [ CO2 ] and warmer temperatures are species-specific ( Engel et al. 2005, Lukac et Al. 2010 ) . This implies the possible to change species composing, competitory ability and natural distribution of tree species in the hereafter ( Saxe et al. 2001 ) . Questions that arise from such possible alterations include whether congenerous black spruce and white spruce would react otherwise to the synergistic effects of dirt heating and fertilization under current and future [ CO2 ] and whether the response would different when the species are grown stray and together in mixture with one another. Black spruce can turn good on both rich highland sites and nutrient-poor peatlands while white spruce merely grown good on fertile highland sites. Black spruce therefore appears physiologically more fictile than white spruce ( Patterson et al. 1997 ) . In response to alimentary demand, black spruce appears to hold lower alimentary demand and wider temperature scope than white spruce ( Viereck and Johnston 1990, Nienstaedt and Zasada 1990, Patterson et Al. 1997, Way and Sage 2008 ) . While black spruce may turn on broad of scope sites, responses of workss to increased atmospheric [ CO2 ] vary with the alimentary malleability of the species ( Brown and Higginbotham 1986 ) . And species happening on alimentary hapless dirts such as black spruce may hold slower response to increased [ CO2 ] and increased alimentary supply ( higher NUE ) than white spruce which normally grows on richer dirts. Furthermore, on fertile sites, fast turning species have high alimentary consumption dynamicss ( Jackson et al. 1990 ) and are able to consume rich alimentary spots before slow turning species with low uptake dynamicss are able to pull out adequate foods ( Jackson et al. 1990, Caldwell et Al. 1996, Lambers et Al. 2008 ) . Plants with high uptake dynamicss may nevertheless be disadvantaged when alimentary supply is low because of greater energy demands ( Jackson et al. 1990 ) . As such, species adapted to low dirt birthrate with lower consumption dynamicss can out-compete species adapted to high alimentary supply, on hapless sites ( Aerts and Chapin 1999 ) . It is hence expected that white spruce will profit more from the increased [ CO2 ] and higher food supply than black spruce. At the low food intervention nevertheless, black spruce may hold higher photosynthetic additions than white spruce. Because the two species have different food demands, alimentary use would be maximised when the two species are grown together. As such, photosynthetic rate in the mixed-species experiment would be higher than in the glandular fever experiments.
Materials and Methods
The experiment was conducted at Lakehead University ‘s Thunder Bay, Ontario Campus, from December 2009-April 2010. One twelvemonth old black spruce ( Picea mariana [ Mill. ] B.S.P. ) and white spruce ( Picea glauca [ Moench ] Voss. ) seedlings raised in a commercial baby’s room were used for the survey. All the seedlings were of unvarying size and signifier at the beginning of intervention. Seedlings of each species were either grown isolated in a individual pot ( single-channel experiment ) or combined together in one larger pot ( mixed-species experiment ) . Soil volume of each seedling in the glandular fever experiment was 1767.38 cm3. For seedlings in the mixed-species experiment, each works was allocated a dirt volume 10 % less than each works received in the single-channel experiment. This was to guarantee that, there is below-ground plant-plant interaction but minimum above land interaction. The turning medium was premium grade vermiculite-peat moss mixture ( 50:50, v/v ) .
Experimental Design and Growth Conditions
Two experiments were carried out at the same time. In experiment one ( mono-grown ) , 8 single seedlings of each species were each grown in a individual pot. In the 2nd experiment ( mixed-species experiment ) , 8 seedlings of each species were grown together in the same pot ( entire of 16 in one pot ) . Apart from the figure of workss per pot, all other conditions were the same for the mono-grown and mixed-species experiments. For each experiment, the design was a split-split-plot with CO2 concentration as the whole secret plan, dirt temperature as the sub-plot and alimentary supply as the sub-sub secret plan. There were two CO2 ( ambient, 360 and elevated, 720 µmol mol-1 ) degrees with two reproductions, two dirt temperature governments within each CO2 intervention. There were two alimentary degrees within each dirt temperature intervention. The last factor, species, was wholly randomised in the design. Agreement of each temperature control box, pot and works within each secret plan was randomised.
The CO2 in the elevated nurseries were generated from electronic ignition natural gas CO2 generators ( theoretical account GEN-2E, Custom Automated Products, Inc, Riverside, CA ) . CO2 degrees in each of the four nurseries were monitored and automatically controlled with Argus CO2 detectors and control system ( Argus, Vancouver, BC, Canada ) . Day-time dirt temperatures were 20 and 25 oC. The 20 oC dirt temperature ( CurrentOT ) represents current optimum for both species ( Grossnickle 2000 ) while the 25 oC ( ProjectedOT ) is based on 5 oC projection. Each day-time dirt temperature was lowered by about 4-6 oC at dark to provide for lower dark temperatures and imitate natural phenomenon. Changing of day-time and night-time dirt temperature occurred 90 proceedingss before air temperature was changed as dirt temperature alteration is slower than air temperature alteration. Each dirt temperature was controlled utilizing a separate control system dwelling of a big leak-proof box ( 112 centimeter broad, 196 centimeter long, 16 centimeter deep ) filled with H2O to a suited degree and a circulative pump ( exemplary AC-2CP-MD, March Mfg. Inc. , Glenview, Illinois, USA ) . The CurrentOT system was equipped with a flow-through cooler-heater ( model 911 ) , while the ProjectedOT system had a flow-through warmer ( theoretical account 210, PolyScience, Nile, Illinois, USA ) to command the H2O to the coveted temperature. The system was insulated to understate heat exchange with the nursery air. The high food government ( 150, 60, 150, 80, 40, 60 ppm N, P, K, Ca, Mg and sulfur severally ) represents fertile boreal sites and was based on Landis ( 1989 ) rapid growing stage alimentary recommendation for conifers. The low food intervention was 10 % of each of the high food intervention concentration. Fertilization was done one time a hebdomad except.
Environment conditions in the nurseries were maintained and controlled utilizing an Argus control system ( Argus, Vancouver, BC, Canada ) as follows: photoperiod of 16 hours, comparative humidness of 55 ± 5 % and twenty-four hours and dark air temperatures of 25 ± 2 oC and 15 ± 2 oC severally. Natural visible radiation in the nursery ( about 700 ?mol m-2s-1 ) was supplemented on cloudy yearss, early forenoons and late eventides with hard-hitting Na lamps. The auxiliary visible radiation was about 220 ?mol m-2s-1 at the canopy degree. Watering was done twice a hebdomad.
Foliar gas exchange was measured 4.5 months into the intervention. Gas exchange was measured on current twelvemonth acerate leafs utilizing PP-Systems Ciras-1 open gas exchange system ( Hitchin, Herefordshire, U. K. ) with a Parkinson conifer foliage cuvette. Environmental conditions in the cuvette were controlled automatically as follows: temperature 25 oC, comparative humidness 50 % , photosynthetically active radiation ( PAR ) 800 µmol m-2 s-1. The visible radiation was supplied from the cuvette ‘s constitutional wolfram lamp. A-Ci curves ( CO2 assimilation rate, A, and sub-stomatal [ CO2 ] , Ci ) were measured consecutive from 50, 150, 250, 360, 500, 700, 900 to 1200 µmol mol-1 [ CO2 ] . Leaf temperature inside the cuvette averaged 24 oC. Net photosynthetic rate ( Pn, µmol CO2 m-2 s-1 ) , stomatous conductance ( gs, mmol m-2 s-1 ) and transpiration rate ( E, mmol H2O m-2 s-1 ) were estimated at growing [ CO2 ] based on Farquhar et Al. ( 1980 ) . Photosynthesis was besides measured at a common ambient, 360 µmol mol-1 [ CO2 ] ( Pn360 ) .
Photosynthetic H2O usage efficiency ( WUE, µmol CO2 mmol H2O-1 ) , sum of C addition per unit H2O was calculated as: WUE = Pn/E, where Pn and E were rates at growing [ CO2 ] . Maximal carboxylation capacity of Rubisco ( Vcmax ) , maximum photosynthetic negatron conveyance capacity ( Jmax ) and triose phosphate use ( TPU ) were estimated from the A-Ci curves utilizing an A-Ci based on Sharkey et Al. ( 2007 ) . All gas exchange parametric quantities and Rubisco biochemical activities are expressed on a projected leaf country footing. Leaf country was determined utilizing the Regent Winseedle system ( Regent Instruments, Qu & A ; eacute ; bec City, QC, Canada ) .