Master’s thesis in Agricultural Development (45 ECTS) Title Morphological and physiological responses of halophytes: Atriplex halimus, A. lentiformis, A. nummularia, Amaranthus caudatus and Chenopodium quinoa Willd. to increasing levels of salinity Author Victoria K. S. Munkager Academic advisor: Sven-Erik Jacobsen Academic co. supervisor: Yaosheng Wang Submitted: 15/01/15 Preface This master’s thesis has been conducted under The Department of Plant and Environmental Sciences, Faculty of Science, University of Copenhagen (KU) and the International Center for Agricultural Research in Dry Areas (ICARDA) under the program Diversification and Sustainable Intensification of Production Systems (DSIPS) as the finalizing component in achieving my degree as Cand. Scient in Agricultural development. 3 months were spend in Amman, Jordan conducting field work as a mandatory element of a master’s thesis in Agricultural Development. The work was partially funded by scholarships through University of Copenhagen and my supervisor Sven-Erik Jacobsen. The DSIPS program funded the entire field work conducted in Jordan. Images of leaves were processed by Sawsan Hassan at ICARDA to provide estimates of leaf area. The analyses of soil and water were performed by the NCARE laboratory. The remaining data collection and analysis is my original work. Victoria K. S. Munkager 15.01.2015 i Abstract 7 % of the world’s agricultural lands are adversely affected by salinity, a state that makes them less capable of supporting agricultural systems. Assessing growth and physiological mechanisms associated with high tolerance in halophytes may provide useful tools in understanding and breeding for salinity tolerance as well as assisting in providing alternative underutilized candidates for reclamation and remediation of saline soils. This study evaluated the salinity tolerance of five halophytes: Atriplex halimus, A. lentiformis, A. nummularia, Amaranthus caudatus and Chenopodium quinoa Willd. variety titicaca exposed to moderate to severe levels of salinity. 25 days after sowing, seedlings were exposed to 5 different levels of salinity with electrical conductivity values of the saturated paste ranging from 8.7 dS/m to 105.8 dS/m. After approx. eight weeks of salinity exposure growth, plant water relations, water use efficiency, stomatal resistance, chlorophyll content and total leaf Na+ and K+ were measured. None of the five species survived more than 23 days of salinity exposure at the two highest salinity levels. Although significant growth reductions in response to increasing salinity were observed for all species there were clear between-species distinctions to be made pertaining to the degree of salinity stress experienced by the plants and which underlying mechanisms were mostly affected. Amaranth and quinoa were found to be Na+ includers consequently altering the ratio between leaf K+/Na+ and inducing premature leaf senescence due to ionic stress. Contrastingly, all three atriplex species were found to be Na+ excluders indicating that control of Na+ uptake and transport plays an essential role in the salinity tolerance of atriplex. The stomatal resistance was adversely affected in nummularia, amaranth and quinoa and to some extent in halimus and lentiformis indicating that increasing salinity provokes a water conserving strategy in the plants which resulted in either unaltered or improved water status in all of the species. Reductions in chlorophyll content in response to salinity were observed in some but not all of the species and it is hypothesize that in quinoa, a C3, and amaranth, a C3-C4, species chlorophyll content reductions lower the risk of photoinhibition under salinity- induced stomatal-limiting photosynthesis inhibition. Overall it can be concluded the significant differences in plant response, both morphological and physiological, to salinity exists, not only between species, but also within the same species over a salinity exposure gradient of EC: 8.7-26.8 dS/m. ii Resumé 7 % af verdens landbrugsarealer kan karakteriseres som saltholdige. Høje koncentrationer af salte resulterer i en reduktion af jordens overordnede produktivitet grundet et ufavorabelt osmotisk potentiale og høje salt koncentrationer. Ved at undersøge hvordan vækst og underliggende fysiologiske mekanismer påvirkes af at vokse i saltholdige jorde kan man komme tættere på at forstå individuelle arters ydeevne samt hvilke mekanismer der er ansvarlige for salttolerance. Denne forståelse kan være essentiel viden for planteforædling af mere modstandsdygtige arter samt til at klarlægge om arter kan anvendes til at forbedre produktiviteten af saltholdige jorde der ellers ikke kan understøtte væksten af saltsensitive arter. I dette eksperiment blev fem forskellige saltelskende arter udsat for moderat til ekstremt saltholdige jorde. Arterne i forsøget var: Atriplex halimus, A. lentiformis, A. nummularia, Amaranthus caudatus og Chenopodium quinoa Willd. varietet titicaca. 25 dage efter såning blev plantespirerne udsat for fem forskellige grader a saltstress, hvoraf den laveste behandling havde en elektrisk ledningsevne på 8.7 dS/m og den højeste 105.8 dS/m. Efter 8 uger med saltstress blev plantevækst, vandindhold, vandudnyttelseseffektivitet, stomatal konduktans, klorofyl indhold og bladkoncentration af Na+ og K+ målt. Ingen af de fem arter overlevede længere end 23 dage i de to mest saltholdige behandlinger. Plantevæksten blev betydeligt reduceret som et resultat af øget saltstress for alle fem arter, men analyserne afslørede derudover at der er forskel mellem arter i grad af påvirkning samt hvilke fysiologiske parametre der påvirkes mest. Amarant og quinoa viste tydelige tegn på akkumulation af Na+ i deres blade i takt med saltstress, hvilket resulterede i at forholdet mellem K+/Na+ blev reduceret og at individuelle blade viste tegn på tidlig ældning. Derimod var indholdet af Na+ uændret af graden af saltstress hos de tre atriplex arter hvilket indikerer at disse arter har effektive mekanismer til at styre optag og bevægelse af Na+ i planten. Stomatal konduktans var negativt påvirket af salt for nummularia, amarant, quinoa og i mindre grad halimus og lentiformis, hvilket signalerer at saltstress giver planterne incitament til at konservere vand ved at lukke stomata. Effektiviteten af denne mekanisme var tydelig at se på det uændrede eller forbedrede vandindhold i planterne. Indholdet af klorofyl i planternes blade blev kun reduceret i nogle af arter. En mulig hypotese for forholdet mellem kloropfyl og stomatal konduktans er, at quinoa og amarant, som ikke er C4 arter, har en eller flere mekanismer, der regulerer klorofylindholdet i forhold til graden af stomatalbegrænsende fotosynteseinhibering. Overordnet set er det muligt a konkludere at der findes betydelige forskelle mellem de forskellige arters reaktion på saltstress, men at der også hersker forskelle indenfor de individuelle arter i takt med at graden af saltstress stiger fra 8.7 til 26.8 dS/m. iii List of abbreviations C3 Photosynthetic pathway in which the first product of CO2 fixation is a 3-carbon intermediate C4 Photosynthetic pathway in which the first product of CO2 fixation is a 4-carbon intermediate CCAFS Climate Change, Agriculture and Food Security CCI Chlorophyll content index CGIAR Consultative Group on International Agricultural Research DAS Days after sowing DAT Days after treatment exposure DSIPS Diversification and Sustainable Intensification of Production Systems DW Dry weight DVCT Digital Vegetation Charting Technique E Transpiration EC Electrical conductivity ECe Electrical conductivity of a saturated paste extract ET Evapotranspiration FW Fresh weight ICARDA International Center for Agricultural Research in the Dry Areas NCARE National Center for Agricultural Research and Extension rs Stomatal resistance iv Table of contents Preface ............................................................................................................................................................... i Abstract ............................................................................................................................................................ ii Resumé ............................................................................................................................................................ iii List of abbreviations ....................................................................................................................................... iv 1. Introduction ................................................................................................................................................. 1 1.1 Salinization, definitions and drivers .................................................................................................. 1 1.2 Plant response to salinity ................................................................................................................... 2 1.2.1 Stomatal resistance .............................................................................................................................. 2 1.2.2 Chlorophyll content ....................................................................................................................... 3 1.2.3 Water use efficiency ...................................................................................................................... 4 1.2.4 Na+ ................................................................................................................................................