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Environmental Research Journal Volume 6, Number 2

ISSN: 1935-3049 © 2012 Nova Science Publishers, Inc.

ECO-PHYSIOLOGICAL RESEARCH FOR BREEDING IMPROVED CASSAVA CULTIVARS IN FAVORABLE AND STRESSFUL ENVIRONMENTS IN TROPICAL/SUBTROPICAL BIO-SYSTEMS Mabrouk A. El-Sharkawy1, Sara M. de Tafur2 and Yamel Lopez2 1

Centro Internacional de Agricultura Tropical (CIAT) International Center for Tropical Agriculture, Cali, Colombia 2 Universidad Nacional de Colombia–Palmira, Colombia

ABSTRACT This chapter highlights eco-physiological, breeding and agronomical research on the tropical starchy root crop cassava (Manihot esculenta Crantz) conducted at CIAT. The objectives were developing improved technologies needed to enhance productivity, root quality for human consumption and other uses, as well as conserving natural resources in different tropical/subtropical bio-systems where the crop is grown. Laboratory and field studies have elucidated several physiological/biochemical mechanisms and plant traits underlying productivity in favorable conditions and tolerance to stressful environments such as prolonged water stress and marginal low-fertility soils. Cassava is endowed with inherent high photosynthetic capacity expressed in near optimal environments that correlates with biological productivity and storage root yield across environments and wide range of germplasm. Long leaf life, and hence better leaf retention, with reasonable photosynthetic rates was also associated with high yields, particularly in prolonged drought conditions. Extensive fine rooting systems capable of exploring deeper wet soils is another mechanism enhancing tolerance to water stress coupled with stomatal sensitivity to both atmospheric humidity and soil water shortages. Cassava leaves possess elevated activities of the C4 phosphoenolpyruvate carboxylase ( PEPC) enzyme that correlate with photosynthesis and productivity in field-grown crops, indicating its importance as another selectable trait particularly for stressful environments. When cassava recovers from stress, it rapidly forms new leaves with higher photosynthetic capacity, which compensates for yield reductions from the previous prolonged stress. Selecting for medium-statured and short cassava instead of tall cassava is advantageous for saving on nutrient uptake and ensuring higher nutrient-use efficiency for root production without sacrificing potential yield. Germplasm from the core collection was screened for tolerance of soils low in P and K, resulting in the identification of several accessions with good levels of tolerance. In cooler zones such as higher altitudes in the 

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Mabrouk A. El-Sharkawy, Sara M. de Tafur and Yamel Lopez tropics and lowland subtropics, cassava growth is slower and the crop stays in the ground for longer time to achieve adequate yields. Under these conditions, leaf formation is slower, leaf photosynthesis is much reduced, but leaf life is longer. Wide genetic variations exist for photosynthesis that may be valuable for selecting and breeding for genotypes that better tolerate cool climates. Combining enhanced leaf photosynthesis with the normally longer leaf life in cool climates may improve productivity. Results also point to the importance of field research versus greenhouse or growth-chamber studies that do not calibrate for representative environments to account for acclimation factors. Calibration becomes even more critical when data from indoor-grown plants are used to extrapolate to the field or to develop crop models. Basic research can be cost-effective, with high returns, even if slower. It can be especially successful when conducted in collaboration with multidisciplinary and/or multi-institutional teams that follow wellplanned strategies and are focused towards fulfilling a set of high priority goals and objectives. Much remains to be done to further improve productivity while conserving dwindling natural resources such as water and land in view of the observed global climate changes. Developing countries, in particular, need more support to continue with maintenance research, which aims to upgrade previous findings and technologies; contribute to sustainable agricultural, economic, and social developments; and enhance food supply to meet increasing demands.

INTRODUCTION Cassava (manioc, yuca, or mandioca; Manihot esculenta Crantz, Euphorbiaceae) is a perennial shrub of the New World and became an important both as a cash crop and a subsistence crop of resource-limited farmers in Africa, Asia, and Latin America and the Caribbean with world production of more than 200 million tons of storage fresh roots annually (El-Sharkawy, 1993) (Figure 1). The plant is outbreeding heterozygous polyploidy species possessing 2n =36 chromosomes. The germplasm is genetically diverse due to botanical, social and environmental influences. Botanically, the species is monoecious (female and male flowers are separated on the same branch) that enhances percentage of natural outcrossing (Jennings and Iglesias, 2002). Socially, man exerted great influence on the diversity of the species since its domestication more than 6000 years ago in the Amazonian region of Brazil (Allem, 2002). Women in particular played a significant role in increasing the genetic diversity of cassava where it is grown. For example, in Africa where newly-wed women move to their husbands’ villages after marriage, they bring with them planting materials of their-choice of cassava varieties from their home farming communities (Delêtre et al., 2011). Environmentally, natural hybridization is a dynamic process and an important driver for genetic diversity where farmers keep mixed cultivars on their fields that normally results in the appearance of new races, some of which are targets for selection and adoption for their desired traits. The storage roots are used as a source of carbohydrate (eg, 80-87% starch of dry matter) as protein is less than 3 percent in dry roots. They are grown mainly for human consumption, either prepared fresh as in the case of sweet cultivars low in cyanogenic glucosides, or after processing into dry products such as flour, starch and animal feed in the case of bitter cultivars high in cyanogenic glucosides (Balagopalan, 2002; Durfour, 1988; Essers, 1995; Westby, 2002; Montagnac et al., 2009a,b).

Eco-Physiological Research for Breeding Improved Cassava Cultivars …

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Figure 1. Clockwise: mature cassava plants, young cassava plantation, and samples of storage roots. Source: CIAT; IITA; Wikimedia Commons, David Monniaux.

Cassava leaves are also consumed and constitute an excellent source for protein supplement (leaf crude protein contents on a dry basis range among cultivars from 21% to 39%; Yeoh and Chew, 1976; Ravindra, 1993), minerals and vitamins for the human diet in many African and Asian countries, as well as in certain regions of Brazil (Lancaster and Brooks, 1983; Montagnac et al., 2009b; Djuikwo et al., 2011). Nevertheless, cassava roots and leaves are deficient in sulphur-containing amino acids (eg, methionine and cysteine; Gil and Buitrago, 2002, and Table 1). Its often poorly-processed food products might contain some anti-nutrient elements such as free HCN, phytates and polyphenols, and particularly acetone cyanohydrin, which is commonly associated with an upper motor neuron disease known as ´konzo syndrome´ in some African countries (Cliff et al., 2011; Tylleskar et al., 1992; Tylleskar, 1994). This occurs mainly with large intake of inadequately processed bitter-cassava products in areas hit by long drought and with shortages of balanced diets.

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Mabrouk A. El-Sharkawy, Sara M. de Tafur and Yamel Lopez

Table 1. Contents of some amino acids in cassava storage roots and leaves, based on dry weight and total protein (adapted with modifications after Gil and Buitrago, 2002). Note the low contents of cysteine and methionine Amino acid

Storage roots % of dry wt.

% of protein

Leaves% of dry wt.

% of protein

Alanine

0.15

5.70

1.70

6.10

Arginine Aspartic acid Cysteine

0.29 0.13 0.01

11.00 4.90 0.40

1.48 2.44 0.21

5.30 8.70 0.70

Glutamic acid

0.15

5.70

1.99

7.10

Glycine Histidine

0.01 0.07

0.40 2.60

1.73 0.66

6.20 2.30

Leucine

0.31

11.30

2.72

9.70

Lysine Methionine

0.07 0.03

2.60 1.00

1.87 0.36

6.70 1.30

Phenylalanine Serine

0.03 0.04

1.00 1.50

0.92 1.68

3.30 6.00

Threonine

0.03

1.00

1.35

4.80

Tyrosine

0.01

0.40

0.89

3.20

Some of those compounds may also act as anti-oxidants and anti-carcinogens, which may result in adverse effects on health if ingested in large amounts (Montagnac et al., 2009a,b; Tsumbu et al., 2011). The crop also has many different alternative uses as processed food, animal feed, starch, alcohol and biofuel. Compared to other energy crops, cassava under favorable production conditions produces more annual ethanol per unit land area (Jansson et al., 2009; Sierra et al., 2010, Table 2) [but also see other estimates (Johnston et al., 2009; Spiertz and Ewert, 2009) ]. Another value-added potential is the possibility of genetically engineer cassava toward transfer and overexpressing the genes controlling ´human serum albumin´ protein (HSA) in cassava storage roots that might lead to economic production of highly needed HAS (current demand is about 500 or more tons annually), an achievement recently obtained with rice grains by Chinese researchers (He et al., 2011). In contrast to water-loving rice, cassava is a versatile and resilient plant that can tolerate and produce reasonably well in harsh environments under hot weather coupled with prolonged drought and marginal soils. Water requirements for the production of one ton of dry matter in cassava storage roots and in rice grains are approximately 400-500 and 35004000 tons, respectively. In view of shortages in irrigation water resources and/or in rainfall that will probably be further aggravated in most tropical/subtropical regions, caused by predicted Global Climate Change when average temperature of earth atmosphere reaches probably 2.4-6.5 ºC above current by the end of this century (El-Sharkawy and De Tafur, 2011), the comparative advantage would be at the side of cassava.


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Table 2. Comparative advantage of cassava as a potential biofuel crop versus other energy crops (adapted with modifications after Jansson et al., 2009). Yield estimates are based on favorable production conditions Crop species

Average yield (Ton ha-1 year-1)

Conversion rate to bioethanol (Liter ton-1)

Bioethanol yield (Liter ha-1 year-1)

Cassava

40 (fresh storage roots)

150

6,000

Sugarcane

70 (stalks)

70

4,900

Sweet sorghum

35 (biomass)

80

2,800

Rice

5 (grains)

450

2,250

Maize

5 (grains)

410

2,050

Wheat

4 (grains)

390

1,560

As countries develop their demand for these products increases dramatically. Under the marginal tropical/subtropical conditions where it is normally grown (with prolonged drought and low-fertility soils), it produces more energy per unit area than other crops, and is highly compatible with many types of intercrop. Thus, cassava’s resilience makes it ideal for food security in marginal environments where other staple crops such as cereals and grain legumes might fail to produce. Because of its metabolic efficiency and other traits described below, cassava is an outstanding starch biosynthesis plant in both lowland subhumid and humid high-productivity environments. It is practically axiomatic that where there’s cassava there’s no famine. It can be grown with minimal inputs, but gives considerably higher yields with fertilizers and good management. The crop is flexible as to time of harvest and can be stored naturally for long periods by keeping the plants in the field with the roots in the soil. Cassava roots are highly perishable once harvested (van Oirschot et al, 2000), and roots must be used immediately or processed into dry products, although pruning three weeks before harvest will reduce deterioration. Therefore, cassava needs to be processed near production fields, making it an ideal vehicle for rural development through creating employment opportunities in the areas where it is grown. Although cassava presumably originated in hot humid climates in the Amazonian lax forests (Allem, 2002), it shows a high degree of tolerance to prolonged drought in areas with low and erratic precipitation of less than 600mm annually, coupled with dry air and high temperatures (hence, high potential evapotranspiration), low fertility soils and high pest and disease pressure such as in Northeastern Brazil, the northern coast of Colombia, the west coast of Peru, some areas in sub-Saharan African countries, and parts of Thailand (ElSharkawy, 1993). Cassava is widely grown in countries of tropical and subtropical Africa, Asia and Latin America between latitudes 30o N and S, from sea level to above 2000 masl by resource-limited smallholder farmers on marginal and highly-eroded low-fertility acidic soils, virtually without the application of agrochemicals (El-Sharkawy, 1993, 2004; Ruppenthal et al, 1997). By not having specific water-stress sensitive growth stages beyond crop establishment (compared with, e.g., grain crops), cassava can survive and produce under conditions where

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other staple food crops, such as grain cereals and legumes, would rarely produce. These inherent characteristics have motivated the expansion of the crop into more marginal lands across many parts of Africa, Asia and Latin America. Notable is its recent expansion in subSaharan Africa (Romanoff and Lynam, 1992) where its tolerance to various edapho-climatic stresses gives an advantage over other staples (Cadavid et al, 1998; De Tafur et al, 1997b; ElSharkawy, 1993; Flörchinger et al, 2000; Fermont 2009). Field research (El-Sharkawy, 2004, 2006a,b, 2007,2009, 2010; El-Sharkawy and De Tafur, 2007, El-Sharkawy et al.,1990,1992a, 1993, 2008) has elucidated several physiological plant mechanisms underlying cassava’s potentially high productivity under favourable hothumid environments in the tropics. Most significant is its inherent capacity to assimilate carbon in near optimum environments, correlating with both biological productivity and storage root yield in a wide range of accessions from cassava core germplasm at CIAT grown in diverse environments over years. Cassava leaves possess elevated activity of the C4 phosphoenolpyruvate carboxylase (PEPC) enzyme that correlates with leaf photosynthesis, leaf photosynthetic nitrogen use efficiency (PNUE) and storage root yield in field-grown plants, indicating the importance of selection for high photosynthesis. Under certain conditions, leaves exhibit a very interesting photosynthetic C3-C4 intermediate behaviour which may have important implications in future selection efforts. In addition to leaf photosynthesis, yield is also correlated with seasonal mean leaf area index (LAI), ie, leaf area duration. Under prolonged water shortage in seasonally dry and semi-arid zones (El-Sharkawy, 1993, 2006a,b, 2007, 2010; De Tafur et al.1997b) once established, the crop tolerates the stress and produces reasonably well compared to other food crops. For example, in environments with less than 700mm of annual rain, with 6 month dryness, improved cassava cultivars can yield over 3t ha−1 of oven-dried storage roots. The underlying mechanisms for such tolerance include stomatal sensitivity to atmospheric and edaphic water deficits, coupled with deeper rooting capacities that prevent severe leaf dehydration (ie, a stress avoidance mechanism), and reductions in the leaf canopy with reasonable photosynthesis over the leaf lifespan. Another stress-mitigating plant trait includes the capacity to recover from stress once water becomes available, by forming new leaves with even higher photosynthetic rates than those of non-stressed crops. Under extended stress (El-Sharkawy and Cock, 1987b; El-Sharkawy and Cadavid, 2002; El-Sharkawy et al., 1992b) the reduction in biomass is greater in the shoot than in the storage root, resulting in a higher harvest index (HI). Cassava conserves water by slowly depleting available water from deep soil layers, leading to higher water-use efficiency (WUE) and nutrient-use efficiency (NUE). In dry environments, leaf area duration and resistance to pests and diseases are critical for sustainable yields. Cassava can survive in the semi-arid zones but a second wet cycle is required to achieve higher yields in this situation, where selection and breeding for early bulking and for medium/short-stemmed cultivars will be advantageous. When grown in cooler zones such as in the tropics at high altitudes and in lowland subtropics, growth is slower and leaf photosynthesis is greatly reduced. Thus, the crop requires a longer period for reasonable productivity. Here, there is a need to select and breed for more cold-tolerant genotypes. This chapter reviews and highlights some of the ecophysiological research conducted at CIAT, particularly under relevant field conditions where most cassava is grown, in relation to breeding improved cultivars for both favorable and stressful environments. The research had


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laid the foundations for cassava breeding and selection of adaptable cultivars under both environments.

CASSAVA TOLERANCE TO DROUGHT AND CONTROLLING PHYSIOLOGICAL MECHANISMS Responses to Extended Water Deficits in the Field Inherent plant mechanisms that may underlie tolerance to drought in cassava have been elucidated and described (El-Sharkawy, 1993, 2004, 2006a,b, 2007, 2010; Caýon et al, 1997; De Tafur et al, 1997a,b; El-Sharkawy et al, 1992b). Most notable is the striking sensitivity of cassava to both changes in atmospheric humidity and soil water deficits – via partially closing its stomata and restricting water loss once exposed to dry air and/or dry soils, thus protecting the leaf from severe dehydration – coupled with leaf capability to partially retain its photosynthetic capacities under prolonged water shortages. Compared to several woody and herbaceous species, cassava is more sensitive to changes in air humidity (Cock and ElSharkawy, 1988b; El-Sharkawy and Cock, 1986; El-Sharkawy et al, 1984d, 1985), and the response has been related to stomatal density and maximum leaf conductance (El-Sharkawy and Cock, 1986; El-Sharkawy et al, 1984d,1985). Cassava leaves possess large number of stomata on the abaxial epidermis (> 400 stomata mm−2; Connor and Palta, 1981; El-Sharkawy et al, 1984a; Guzman 1989; Pereira, 1977) that may underlie the strong response to humidity (El-Sharkawy and Cock 1986; El-Sharkawy et al, 1985). The phenomenon of direct stomatal response to air humidity was observed as early as the late 19th and early 20th centuries by botanists (El-Sharkawy and Cock, 1986; Haberlandt, 1914; Thoday, 1938). Numerous more recent reports have shown that several herbaceous and woody plant species tend to close their stomata in response to dry air, whether observed within the plant community or in attached leaves and isolated epidermal strips (eg, Aston, 1976; Bongi et al, 1987; Bunce, 1981, 1982, 1984; Fanjul and Jones, 1982; Farquhar et al, 1980; Gollan et al, 1985; Hall and Hoffman, 1976; Hall and Schulze, 1980; Held, 1991; Hirasawa et al, 1988; Hoffman and Rawlins, 1971; Hoffman et al, 1971; Jarvis, 1980; Jarvis and McNaughton, 1986; Kappen and Haeger, 1991; Kaufmann, 1982; Körner, 1985; Körner and Bannister, 1985; Lange et al, 1971; Leverenz, 1981; Lösch, 1977, 1979; Lösch and Schenk, 1978; Lösch and Tenhunen, 1981; Ludlow and Ibaraki, 1979; Meinzer, 1982; Pettigrew et al, 1990; Rawson et al, 1977; Sheriff and Kaye, 1977; Schulze, 1986; Schulze and Hall, 1982; Schulze et al, 1972; Tazaki et al, 1980; Tibbitts, 1979; Tinoco-Ojanguren and Pearcy, 1993; Ward and Bunce, 1986). This apparently widespread phenomenon indicates the need for detailed studies and for its consideration while modelling plant community/environment ecosystems (Jarvis and McNaughton, 1986). The adaptive ‘stress avoidance mechanism in cassava’ operating via stomatal sensitivity to both edaphic and atmospheric water deficits is of paramount importance for the crop’s tolerance to prolonged drought (more than three months) combined with hot dry air in seasonally dry and semi-arid zones (De Tafur et al. , 1997a,b; El-Sharkawy, 1993). Coupled with this mechanism is the deeper rooting system (below two metres soil depth) that allows the crop to extract storage water when available (Cadavid et al, 1998; CIAT, 1983, 1994;

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Connor et al, 1981; De Tafur et al, 1997a; El-Sharkawy and Cock, 1987b; El-Sharkawy et al, 1992b; Porto, 1983). Although cassava has sparse fine root systems compared with other crops such as cereals and tropical grasses (Tscherning et al, 1995), it is capable of penetrating into deeper soil layers. This enables the plant to endure long periods of drought, only slowly depleting deeper stored water, with the end results of higher seasonal crop WUE, albeit at reduced productivity (Connor et al, 1981; El-Sharkawy, 1993, 2004; El-Sharkawy and Cock, 1986, 1987b; El-Sharkawy et al, 1992b). Another characteristic of great importance in conserving water under extended stress is the large reduction in light interception (Figure 2; CIAT, 1991, 1992, 1993,1994, 1995) achieved through a reduced leaf canopy due mainly to restricted formation of new leaves, a smaller leaf size, and leaf fall (Connor and Cock, 1981; El-Sharkawy and Cock, 1987b; ElSharkawy et al, 1992b; Palta, 1984; Porto, 1983). Although a reduction in leaf area conserves water, it would also lead to a reduction in total biomass and yield ( CIAT, 1991, 1992, 1993, 1994,1995; Connor and Cock, 1981; Connor et al, 1981; El-Sharkawy and Cadavid, 2002; ElSharkawy and Cock, 1987b, El-Sharkawy et al, 1992b, 1998b; Porto, 1983). However, cassava can recover rapidly once released from stress by forming new leaves.

Figure 2. Light interception in water-stressed and well-watered cassava crops. Measurements were made with light sensors placed on top of the canopy and at soil level in the middle of plots. Note the large reductions in light interception overtime in the stressed crops due to reduced leaf formation, the small size of new leaves and the loss of old leaves, and the increase in light interception after recovery from stress due to the formation of new leaves. Source: CIAT, 1991; El-Sharkawy, 1993.

These increase light interception and canopy photosynthesis, thus compensating for previous losses in biomass, particularly root yield (CIAT 1991, 1992, 1993, 1994, 1995, 1998b; El-Sharkawy, 1993; El-Sharkawy and Cadavid, 2002; El-Sharkawy and Cock, 1987b;


151

El-Sharkawy et al, 1992b). Cassava leaves also remain reasonably active during water shortages in the field (Figure 3). Stressed leaves are capable of maintaining photosynthetic rates around 40–60 percent of that of non-stressed leaves during an entire three month stress period. There are differences among cultivars, with the hybrid CM 489-1 showing the least reduction.

Figure 3. Response of cassava leaf photosynthesis to prolonged water stress (120 d), imposed at 60 d after planting (control, open symbols; stress, solid symbols). From CIAT (1987-1989) Report; El-Sharkawy., 2010.

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Upon recovery from stress, the previously stressed leaves can approach the photosynthetic rates of non-stressed controls. Furthermore, the newly-formed leaves of the previously stressed crop show even higher photosynthetic rates than non-stressed plants (Figure 4). These higher photosynthesis rates coincide with higher stomatal conductance to water vapor, higher mesophyll conductance to CO2 diffusion and higher levels of nitrogen, phosphorus, calcium and magnesium in leaves compared with controls (Cayón et al, 1997; CIAT, 1990; El-Sharkawy, 1993). Moreover, Cayón et al (1997) reported greater mobilization of potassium out of newly developed leaves, with new leaves in previously stressed crops consistently containing an average of 0.79 percent potassium compared with 0.96 percent in non-stressed ones. This suggests a higher demand for assimilates in storage roots, since potassium is normally translocated along with sugars to sinks (Giaquinta, 1983).

Figure 4. Response of field-grown cassava to prolonged midseason water stress. Cassava plants were deprived of water by covering the soil surface with white plastic sheets for three months, commencing 90 d after planting. Plants recovered from stress (see arrows in A and B) under rainfall and supplemental irrigation for the rest of the growth cycle. The control plants received normal rainfall plus supplemental irrigation. Leaf gas exchange was monitored with a portable infrared gas analyzer throughout the stress and recovery periods. A. Leaf photosynthesis during stress and recovery for leaves developed under stress. B. Stomatal conductance. C. and D. Photosynthesis and stomatal conductance for new leaves developed after stress was terminated. From CIAT, Cassava Physiology Section database (1991) Annual Report; El-Sharkawy, 1993.

Accordingly, leaf photosynthesis is also controlled in this case by sink demand and strength (Burt, 1964; El-Sharkawy, 2004; Herold, 1980; Ho, 1988; Humphries, 1967; Nösberger and Humphries, 1965; Pellet and El-Sharkawy, 1994; Thorne and Evans, 1964;


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Wardlaw, 1990), suggesting that the dynamics of potassium in leaves of field-grown cassava might be used as an indicator of root sink strength and source-sink relationships.

Lack of Osmoregulation in Cassava under Extended Water Stress In the same studies, predawn leaf water potential (Figure 5 A,CIAT, 1992) remained around -0.5MPa for all cultivars throughout most of the stress period of three months, with virtually no differences between the stressed and unstressed crops. Midday leaf water potential (Figure 5 B) in all cultivars in both stressed and unstressed crops oscillated between −0.6 MPa (when presumably a lower leaf-to-air vapor pressure deficit (VPD) during a wet period coincided with the time of measurement) and −1.6 MPa (at a much higher VPD during dry/sunny periods), with slight reductions often observed in the stressed crops. These values are within the ranges previously reported for cassava under extended periods of soil water shortages in the field (Caýon et al, 1997; Cock et al, 1985; Connor and Palta, 1981; De Tafur et al, 1997a; El-Sharkawy et al, 1992b; Porto, 1983). They are higher than those normally observed in other field crops under stress, indicating that cassava conserves water and prevents extreme leaf dehydration due to its stomatal sensitivity to stress (ie, stress avoidance mechanism). It appears, therefore, that the phenomenon of osmotic adjustment (OA) in mature leaves developed under water stress and other edaphic stresses, as observed in several other field crops (Ackerson and Hebert, 1981; Hsiao, 1973; Hsiao, et al 1976; Jones and Turner, 1978; Morgan, 1984; Turner et al 1978) is not operating in field-grown cassava, since predawn and midday bulk leaf water potential always remained above −0.8 MPa and −2.0 MPa respectively, during prolonged water deficits. Hence, OA is of a little importance as a possible mechanism underlying cassava tolerance to drought. In recent studies with potted greenhouse-grown cassava, Alves (1998) reported that the greatest increases in solutes after few days of water deficit occurred in the youngest and folded (ie, not fully expanded) leaves, with the least increases in mature ones, pointing to little importance of OA in mature leaves. Nevertheless, such studies need to be carried out on field-grown plants if results are to be extrapolated to field conditions, and for obviating acclimation problems (El-Sharkawy, 2005). Since osmoregulation requires investment and accumulation of solutes and assimilates for its development under stresses, McCree (1986) discussed the relative carbon costs involved in the process of OA in sorghum grown under both water deficit and salinity. This author concluded that the metabolic cost of storing photosynthate and using it for OA was less than the cost of converting it to new biomass, although the cost increased slightly under salinity. Under drought, changes in the biosynthesis, content and distribution of plant growth regulators such as abscisic acid (ABA) within plant organs and tissues (particularly in roots, leaves and buds) may play an important role in sensing changes in both soil water and atmospheric humidity, and in controlling stomatal movements, leaf formation and extension, root growth, bud dormancy. They may also be involved in other biological functions such as activation and expression of PEPC, and in possible switching/induction from C3 to crassulacean acid metabolism (CAM) or C4 photosynthesis in some species (Ackerson, 1980; Chapin, 1991; Chu et al, 1990; Davies et al, 1986; Henson, 1984a; 1984b; Huber and Sankhla, 1976; Jones and Mansfield, 1972; Jones et al, 1987; Radin, 1984; Radin et al, 1982;

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Schulze, 1986; Taybi and Cushman, 1999; Turner, 1986; Ueno, 2001; Walton, 1980; Zeevaart and Creelman, 1988; Zeiger, 1983; Zhang and Davies, 1989).

Figure 5. Leaf water potential in water-stressed and well-watered cassava crops during a stress period initiated at 90 days after planting. Measurements were made with a pressure chamber in a field laboratory. Values are means of 5-10 leaves from the upper canopy. A: predawn; B: midday. Note the small differences between the two crops in the predawn and midday water potential and the increases in water potential in the period of high ambient humidity. The predawn and midday levels were above −0.8 and −2.0 MPa, respectively, indicating the striking stomatal control in cassava regardless of soil water status. Source: CIAT, 1991.


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These remarkable physiological responses to mid-season water stress point to cassava’s potential to tolerate prolonged drought and to its resilience and ability to recover from stress when water becomes available, such as in subhumid zones with intermittent dry spells or in seasonally dry zones with well-marked bimodal rainfall distribution. Under these conditions, longer leaf life coupled with resistance to pests and diseases (Byrne et al, 1982; El-Sharkawy 1993; El-Sharkawy et al., 1992b; Lenis et al., 2006) which allows better leaf retention, is advantageous in saving dry matter invested in leaves and in allowing partitioning of excess photosynthates toward storage roots. In semi-arid zones with less than 600mm of rain annually and with periods of water deficits longer than four to five months, such as in Northeastern Brazil, northeastern Colombia and sub-Saharan Africa (De Tafur et al, 1997b; El-Sharkawy, 1993, 2004, 2007), the crop can survive, albeit with greater losses of leaf canopy and less dry matter in the storage roots. In this ecosystem, a second wet cycle is needed to allow recovery of growth and complete filling of roots. Moreover, the use of gene manipulating/transgenic biotechnology coupled with development of molecular markers to facilitate the introgression into cassava cultivars gene(s) controlling ‘exotic’ and specific drought-tolerance related traits such as ‘staygreen’ was recently suggested (Orek et al. 2008; Vanegas, 2008; Zhang et al. 2010) which may accelerate identifying genes controlling leaf retention, particularly in short-stemmed cassava cultivars with small leaf canopy (El-Sharkawy and De Tafur, 2010).Transgenic cassava lines (eg, SAG12-IPT ) that express a cytokinin biosynthesis gene ( IPT ) from a senescence enhanced promoter (SAG12) showed an extended leaf life. Accordingly, several aspects of senescence are delayed in the leaves of these transgenic lines, including chlorophyll degradation, protein degradation and Rubisco reduction. Cassava´s Rubisco activities are very sensitive to water stress (El-Sharkawy, 2004, 2006 b) which might be enhanced via this line of research. It appears that further physiological and agronomical characterizations of these lines also revealed that the induced expression of the gene ÍPT´ affected photosynthesis, sugar allocation and nitrogen partitioning (Zhang et al., 2010). Nevertheless,Vanegas (2008) reported that a transgenic clone ´529-28´that expressed the gene ÍPT´ showed longer leaf life and greater leaf biomass but with reduction in dry root yield indicating a lack of translation into agronomic advantages. However, since whole plant tolerance to water stress is apparently underpinned by several morphological, anatomical, physiological and biochemical mechanisms and traits, transfering/inserting one or few éxotic genes´ might not be the ultimate panacea for drought tolerance in cassava (Bohnert et al., 1996). Such approach needs to be scrutinized through evaluating these transgenic lines for genetic stability, productivity and tolerance to natural prolonged drought at the farm level. A recent obscure report (Okogbenin et al., 2010), and hence given conclusions, based on deficient experimentation schemes and not substantiated with relevant hard data must be seriously questioned concerning its utility for cassava improvement. In contrast, the pioneering ecophysiological research achievements highlighted in this chapter and elsewhere had laid the foundations for the first time of a successful strategy for cassava improvement in both favorable and stressful environments (El-Sharkawy and Cock, 1987b; Cock and ElSharkawy, 1988a,b; El-Sharkawy et al., 1992b; Hershey and Jennings,1992; Iglesias et al., 1995,1996; El-Sharkawy,1993, 2004, 2006a,b, 2010) . Furthermore, to increase the benefitcost ratio of research, it is recommended here to combine the genetic engineering efforts with available sound agronomic, physiological, biochemical and conventional breeding/genetic information on whole plant responses to extended water shortages under field conditions (i.e.,

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holistic approach, El-Sharkawy, 2006a). Ryder (1984) wisely remarked: “I believe very strongly that the technology we call genetic engineering and its attendant sciences and technologies should be pursued. The potential benefits are worth the pursuit. I do not wish to see the support of genetic engineering undertaken, however, on the basis of broad edicts, at the expense of other aspects of plant breeding and its attendant sciences and technologies”. Donors, scientists, and research managers must be aware of the shortcomings of fashion-type (bandwagon!) research projects/programs that serve mostly short-sighted self-interests. Moreover, abiding by scientific ethics must be the rule against violation of intellectual property rights and plagiarism often occurring in science. In this regard, Garfield (1991) wrote on the problem of literature citation violations, misuse and omissions by researchers coining it “bibliographical negligence and citation amnesia” and suggested that authors should sign a pledge or oath that they have done a minimal search of the literature and that to the best of their knowledge there is no other relevant original work not being correctly acknowledged and properly cited (see Gallagher, 2009).

Cassava Photosynthesis and C3-C4 Intermediate Characteristics Underpenning Tolerance to Drought Previous research on cassava photosynthesis demonstrated the importance of elevated activity of the C4 enzyme PEPC that may partially underlie the high photosynthetic capacity in cassava, which was found to be correlated with high productivity across environments, including marginal ones. Table 3. Activity of C4 PEPC in leaf extracts. Values are mean of four leaves ± SD. Note the much higher activity in cassava as compared to beans, a C3 species, and the very high activity in wild Manihot (about 30–40 percent of the activity in maize, a C4 species). In a group of field-grown cultivars under prolonged water stress, net leaf photosynthesis (Pn) was significantly correlated with PEPC activity measured on the same leaves (ElSharkawy, 2004), indicating the importance of selection and breeding for elevated PEPC activity. It is also noteworthy that wild Manihot also possess an additional short palisade layer beneath the lower leaf surface and with numerous stomata on both leaf surfaces, two traits advantageous for enhancing photosynthesis (El-Sharkawy, 2004). Sources: Cassava Physiology Section database; El-Sharkawy, 2004; El-Sharkawy and Cock, 1990; El-Sharkawy, Bernal and López, unpublished observations) PEPC activity (µmol NADH) Species Maize (cv CIMMYT 346) Common beans (cv Calima G4494) Cassava M Mex 59 M Nga 2 Wild species Manihot grahami Manihot rubricaulis

−1

−1

gfw min

mg chl−1min−1

15 ± 1.8 0.2 ± 0.07

7 ± 3.6 0.3 ± 0.1

3.2 ± 0.6 1.3 ± 0.1

2.2 ± 1.0 0.4 ± 1.0

4.0 ± 0.9 5.8 ± 0.6

2.8 ± 1.2 3.4 ± 1.3


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Thus, several cassava cultivars and wild species showed activities from 15 to 25 percent of those in C4 species such as maize and sorghum, with activities ranging from 1.5 to over 5µmol mg-1 chlorophyll min-1 (Table 3; Bernal, 1991; CIAT, 1990, 1991, 1992, 1993, 1994; Cock et al, 1987; De Tafur et al, 1997b; El-Sharkawy 2004, 2009; El-Sharkawy and Cock, 1987a; 1990; El-Sharkawy et al, 1990,1992a ,1993; López et al, 1993; Pellet and ElSharkawy, 1993a). These PEPC activities observed in cassava and its wild relatives are much greater than those observed in C3 species such as field beans, and are comparable to activities found in several C3–C4 intermediate Flaveria species with a limited functional C4 cycle and two to three times greater than those in the C3–C4 Kranz-like Panicum milioides (Brown and Bouton, 1993; Ku et al, 1983).

EFFECTS OF WATER STRESS ON PHOTOSYNTHESIS ENZYMES Under prolonged drought in the field coupled with hot dry air under intense solar radiation, leaves of over 100 cassava cultivars remained photosynthetically active, although at much reduced rates (De Tafur et al, 1997a,b; El-Sharkawy, 1993; El-Sharkawy et al,1990, 1992b). Three weeks after initiation of water stress, PEPC, ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) and the C4 decarboxylase nicotinamide adenine dinucleotide–malic enzyme (NAD–ME) showed reduced activity, with the highest reduction occurring in Rubisco activity in one month-old leaves that had developed before stress started (Table 4; CIAT 1993). The PEPC/Rubisco ratio, which may indicate the relative importance of these two enzymes, was also reduced by stress. However, when leaves that developed under stress for eight weeks were assayed, PEPC activity across all clones was 13 percent greater than unstressed crops with differences among accessions (Table 5; CIAT 1993), whereas Rubisco activity was 42 percent less in the stressed crops. This differential effect of stress on the activities of these two key photosynthetic enzymes resulted in a much higher PEPC/Rubisco ratio in the stressed crops as compared to the unstressed ones. Table 4. Activity of photosynthetic enzymes in leaf extracts of field-grown cassava as affected by three weeks of water stress commencing 92 days after planting (DAP) at Santander de Quilichao, Colombia. Values are means ± SD; activities are expressed in µmol mg chl−1 min−1. Note the much greater reduction in activity of the C3 Rubisco, as compared with the C4 PEPC, in leaves that developed before initiation of stress. Sources: Cassava Physiology Section database; CIAT, 1993; Lopez and El-Sharkawy, unpublished observations Unstressed Clone

PEPC

CM 4013-1 0.37±0.60 CM 4063-6 0.57±0.06 SG 536-1 0.67±0.05 M Col 1505 0.45±0.01 Average 0.51 % change due to stress

Rubisco

NAD-ME

0.31±0.05 3.72±0.11 0.39±0.20 1.18±0.08 1.4

0.40±0.06 0.39±0.06 0.55±0.18 0.16±0.02 0.38

Stressed PEPC/ Rubisco 1.19 0.15 1.72 0.38 0.86

PEPC

Rubisco

NAD-ME

0.31±0.03 0.41±0.04 0.57±0.12 0.49±0.10 0.45 −12

0.41±0.09 1.69±0.10 0.81±0.20 0.49±0.12 0.85 −39

0.19±0.05 0.17±0.09 0.39±0.50 0.29±0.06 0.26 −32

PEPC/ Rubisco 0.76 0.24 0.70 1.00 0.68 −21

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Table 5. Activity of some photosynthetic enzymes in leaf extracts of field-grown cassava as affected by eight weeks of water stress commencing at 92 DAP at Santander de Quilichao; values are means ± SD; activities are expressed in µmol mg chl−1 min−1. Note the large reductions in activity of the C3 Rubisco, as compared to the C4 PEPC, in leaves that developed under stress resulting in a greater PEPC/Rubisco ratio. This photosynthesis-based biochemical assay has proved its utility in selection for tolerance to prolonged drought, see data in Tables 12 and 13. Sources: Cassava Physiology Section database; CIAT, 1993; Lopez and El-Sharkawy, unpublished observations) Unstressed

Stressed

Clone PEPC CM 4013-1 0.86 ± 0.12 CM 4063-6 0.89 ± 0.05 SG 536-1 1.46 ± 0.42 M Col 1505 1.09 ± 0.10 Average 1.08 % change due to stress

Rubisco 0.28 ± 0.10 2.30 ± 0.03 0.44 ± 0.12 0.57 ± 0.13 0.9

PEPC/ Rubisco 3.10 0.39 3.30 1.90 2.2

PEPC

Rubisco

1.18 ± 0.17 1.42 ± 0.26 1.33 ± 0.22 0.96 ± 0.16 1.22 +13

0.30 ± 0.01 0.62 ± 0.02 0.25 ± 0.08 0.89 ± 0.14 0.52 −42

PEPC/ Rubisco 3.9 2.3 5.3 1.1 3.2 +45

These data indicate that, under prolonged water deficit, the relative importance of the C4 PEPC versus the C3 Rubisco becomes more pronounced, and lends support to the hypothesis that the C4 PEPC enzyme may play a significant role in photosynthetic activity under drought coupled with high air temperatures (CIAT, 1993; El-Sharkawy, 2004). This is of paramount importance in reducing both photorespiratory and mitochondrial dark CO2 losses, and in increasing net carbon uptake and hence productivity. Moreover, the recent evidence about the possible location of PEPC in the upper end of the long palisade parenchyma further supports the role of PEPC involvement in refixation/recycling respiratory CO2 when the highly dense abaxial stomata are partially closed under drought, high solar irradiances and high temperatures coupled with dry air. This is particularly the case in hypostomatous leaves that normally possess more than 400 stoma per square mm (El-Sharkawy et al., 1984a). Besides increasing carbon uptake, such a CO2 recycling mechanism protects the leaves from photoinhibition via dissipating excess absorbed photons (Osmond et al, 1980; Osmond and Grace, 1995).

CHARACTERIZATION OF WATER STRESS TOLERANCE Mechanisms Underlying Stomatal Response to Air Humidity Many workers have reviewed and discussed the possible mechanisms underlying stomatal sensing of changes in air humidity and the role of the so-called ‘peristomatal transpiration’ – first hypothesised by Seybold (1961/1962) – due to water loss from the cuticle of the guard and subsidiary cells and their adjacent epidermal cells, in the control of stomatal movement (Lange et al, 1971; Lösch and Schenk, 1978; Lösch and Tenhunen, 1981;Maier-Maercker, 1979a,b, 1983; Meidner, 1976; Meidner and Mansfield, 1968; Sheriff, 1977, 1979, 1984; Tyree and Yianolis, 1980; Zeiger, 1983). Support for Seybold’s hypothesis


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on the role of peristomatal transpiration was demonstrated through extensive research by German workers using intact leaf and isolated epidermal strip systems without water stress (Lange et al, 1971; Maier-Maercker, 1979a,b, 1983; Lösch and Tenhunen, 1981). Others (Meidner and Mansfield, 1968) have argued that stomatal movements are most likely to be affected primarily by the water status of mesophyll tissue through a feedback reaction, rather than by changes in atmospheric humidity. Kramer (1983) cautioned against the proposed role of peristomatal transpiration until more information became available concerning the degree of cutinization of mesophyll tissue (where the bulk of evaporation presumably takes place), of the epidermis and of the inner/external walls of guard cells. Appleby and Davies (1983) demonstrated possible sites of evaporation in cuticle-free areas in the guard cell walls of oak (Quercus robur), poplar (Populus nigra) and Scots pine (Pinus sylvestris) which could be exposed on the outside of the leaf during stomatal closure in dry air. Also, Körner and Cochrane (1985) reported lesser degrees of cutinisation of the external walls of guard cells in Eucalyptus pauciflora that may underlie its stomatal sensitivity to changes in air humidity. Sheriff (1977, 1979, 1984) suggested that the mechanism underlying direct stomatal response to low humidity involves both evaporation from the epidermis and lower hydraulic conductivity within the leaf that may accelerate water stress in the epidermis, regardless of leaf water content. Recently, Pieruschka et al. (2010) reported that the water balance in the epidermis is very sensitive to differences between the transpiration rate and the rate at which absorbed radiation produces water vapor inside the leaf. These authors suggested that leaf heat load is tightly linked to water transport from mesophyll cells, through the epidermis, to the leaf’s environs. This important finding further explains why cassava leaves orient themselves towards the sun in early morning and late afternoon (also called heliotropism or sun tracking) when VPD is lowest, and droop at mid-day (also called paraheliotropism or sun avoidance) when VPD is highest (El-Sharkawy and Cock 1984; Berg et al.1986), thus optimizing water-use efficiency. Tyree and Yanoulis (1980) used physical models of the substomatal cavity to calculate water vapour diffusion patterns and concluded that, due to high evaporation from guard cells, stomata could close in direct response to low humidity. They suggested that localised water stress or dehydration in guard cells may take place due to a high leaf resistance to the flow of liquid water from the minor leaf veins to the guard cells. The strong association between stomatal density (ie, exposed epidermal areas) and the degree of sensitivity to changes in air humidity that was observed in well-watered plants across many herbaceous and woody species (El-Sharkawy et al, 1985) may indicate the occurrence of localised dehydration in the stomatal apparatus and adjacent exposed epidermal cells, hence supporting the role of peristomatal transpiration in controlling stomatal movement. Moreover, the poor physical connection between the numerous stomatal areas (where evaporation may take place) and the mesophyll tissue observed in the cassava leaf (ElSharkawy and Cock, 1986) could accelerate water stress in the epidermis and stomatal apparatus, thus leading to the striking sensitivity to changes in atmospheric humidity without noticeable decreases in bulk leaf water potential (Figures 5, 6, 7, and 8; also see Cayón et al, 1997; Connor and Palta, 1981; De Tafur et al, 1997a; El-Sharkawy, 1990; El-Sharkawy and Cock, 1986; El-Sharkawy et al, 1984d, 1992b; Porto, 1983). This conclusion was further substantiated by the closure of stomata in field-grown cassava in response to high wind speed, despite wet soil and high bulk leaf water potential conditions (El-Sharkawy, 1990).

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Figure 6. Responses of leaf photosynthesis to changes in air humidity. Plants of cv M Col 88 grown in 40 liter pots. The pots were left in the open throughout the growing period and plants were regularly irrigated whenever needed. Pots of unwatered plants were covered at soil level with plastic covers 33 days after planting to exclude rain water. Measurements of gas exchange were conducted on fully expanded young attached leaves under controlled laboratory conditions at saturating photon flux density and normal air using differential multi-channel open end infra red gas analyser. Source: El-Sharkawy and Cock, 1984.

a


161

b

c Figure 7. Respnses of: (a) leaf photosynthesis; (b) transpiration; and (c) leaf conductance to water vapour to stepwise increases in leaf-to-air VPD in cv M Col 90. Growth conditions and gas measurements were as in Figure 6. Source: El-Sharkawy and Cock, 1984.

Bunce (1985) also reported on greater water loss under high wind speed from the outer surface of the epidermis of herbaceous species, providing further evidence in support of peristomatal transpiration.

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Figure 8. (a) Responses of leaf photosynthesis to changes in air humidity in field-grown cassava (cv M Col 1684) with and without misting using high pressure misters installed in the misted plot (about 1000m2) at 5 x 5m distances. Furrow irrigation was applied weekly to both crops to keep the soil wet throughout the trial period. The misted plot (misted from 1000–1500h daily) was guarded by two rows of elephant grass as wind barriers. Gas exchange was conducted by using the syringe method to withdraw air samples from the leaf chamber enclosing upper canopy fully expanded attached leaves exposed to sun radiation greater than 1,000µ mol photons m−2 s−1, the CO2 concentration in samples was determined by using the same gas analyser as in Figure 6 but in a closed system. The gas analyser was periodically calibrated using a series of standard CO2 cylinders with a range of concentrations. Sources: Cock et al, 1985; El-Sharkawy and Cock, 1986. (b) Oven-dried storage root yield in cv M Col 1684 at periodical harvests after 3, 6 and 9 weeks of misting. Ages of plants at harvests were 65, 85 and 105 days, respectively. The differences in yield between the misted and control crops were significant at all harvests (p < 0.01). Top biomass and LAI were not different between the two crops, while total biomass was significantly larger after 6 and 9 weeks of misting. Sources: Cock et al, 1965; El-Sharkawy and Cock, 1986. (c) Response of leaf WUE to VPD in field-grown cassava in a mid-altitude, warm sub-humid climate. Measurements of gas exchange in upper canopy attached leaves were always made between 0900 and 1300h local time, when solar irradiance always exceeded 1,000µ mol photons m−2 s−1, as measured using a portable infrared gas analyser. VPD levels progressively increased from morning to midday. 33 clones were evaluated and were grouped into humid, sub-humid/seasonally-dry and semi-arid habitats. Sensitivity to VPD increased from the humid to the semi-arid habitat. Differences between plant groups illustrate the genetic diversity within cassava germplasm in response to changes in atmospheric humidity. Sources: El-Sharkawy, 2004; El-Sharkawy, Amézquita, Ramirez and Lema, unpublished observations.


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BREEDING STRATEGY AT CIAT FOR DROUGHT TOLERANT CASSAVA CULTIVARS Introduction, Background and Rational There was some limited early work by western colonial communities in parts of Africa, Asia and Latin America during the first half of the 20th century (Cock, 1985; Cours, 1951; Hunt et al, 1977; James, 1959; Nijholt, 1935; Verteuil, 1917, 1918). However, despite its potential productivity, cassava has received little attention from either policy makers or researchers in developing countries where it is widely grown. Although cassava is the most important carbohydrate food source after rice, sugarcane and maize for over 500 million people in developing countries within the tropical and subtropical belt, the crop was overlooked by the so-called ‘Green Revolution’. This early effort by the international agricultural research centres, CIMMYT at Mexico and IRRI at Philippines, in the 1960s focused on improvement of the main cereal crops such as wheat, rice and maize, with the help of international agricultural development and research agencies supported by few western donors. For this reason, the Consultative Group on International Agricultural Research (CGIAR) that was established in 1971 under the sponsorship of the World Bank, the United Nations Development Programme (UNDP), and the Food and Agriculture Organization of the United Nations (FAO; Wortman,1981), gave high priority to research on other crops including cassava, as well as on production ecosystems in the humid tropics of Africa and South America. The work was conducted in Africa through the International Institute of Tropical Agriculture (IITA), located in Nigeria, and in South America through the Centro Internacional de Agricultura Tropical (CIAT) located in Colombia. Given the financial support that became available, core teams of international multidisciplinary scientists were able, for the first time, to conduct extensive research on cassava, in collaboration with the few existing national research programmes. They achieved improvements in germplasm collecting and characterization, breeding, agronomy and cropping systems management, pests and diseases, and utilisation of the crop, based on a better understanding of the physiological processes involved. Recent publications (Hillocks et al, 2002; Kawano, 2003) review results on the many aspects of cassava research in Africa, Asia and Latin America over the last three decades. The multidisciplinary cassava programme at CIAT was established in the early 1970s, with a global mandate for cassava. The programme centred its research strategy on collecting, conserving, and characterizing the germplasm available world-wide that originated mainly in Latin America, and on selecting and breeding more broadly-adapted germplasm for the various environments prevailing in the tropics and subtropics in both Latin America and Asia. In the early stages, breeding objectives were directed toward developing high yielding cultivars under favorable conditions in the absence of biotic and abiotic stresses (Cock et al, 1979; Kawano et al, 1978). This strategy was based mainly on selection for genotypes with high yield per unit land area (in contrast to traditional vigorous cultivars/landraces suited for

164


intercropping), high dry matter content (ie, high starch content) in the storage roots, and a higher HI (expressed as root yield/total plant biomass) than the values of 0.5 or less found in most low-yielding traditional varieties and landraces (Kawano, 1990, 2003). This early work showed that cassava germplasm is genetically diverse with a high potential for productivity under near optimum environments. Thus, the need to transfer traits from wild relatives (reasonably advocated even as recently as 2010 by Nassar and Ortiz) was largely obviated. However, since most cassava production occurs in environments having various degrees of stress, and with little or no production inputs by resource-limited small farmers, the goals of subsequent breeding efforts have centred upon selecting and developing cultivars with reasonable and stable yields, plus the ability to adapt to a wide range of biotic and abiotic stresses (Hershey, 1984; Hershey and Jennings, 1992; Hershey et al, 1988; Jennings and Iglesias, 2002; Kawano, 2003; Kawano et al, 1998). This strategy was further stimulated by the inherent capacity of cassava to tolerate adverse environments, particularly where other main staple food crops such as cereals and grain legumes would fail to produce, and by the opportunity that it offered to reduce or avoid the negative environmental consequences of adopting high-input agrochemically-aided production systems (El-Sharkawy, 1993). The strategy took advantage of the wide genetic diversity within the more than 6,000 accessions, mostly of Latin American origin, conserved at CIAT Headquarters. It also took advantage of the diversity of conditions with high pest and disease pressures that exist in various ecozones throughout Colombia – seven or eight zones with a wide range of climatic and edaphic conditions that are representative of most cassava production ecosystems in the tropics and subtropics (El-Sharkawy, 1993; Hershey and Jennings, 1992). In the light of this environmentally-sound breeding strategy, research on cassava physiology has focused on investigating both basic and applied aspects of the crop under the prevailing environments, mainly in the field, to elucidate and understand better the characteristics and mechanisms underlying productivity and stress tolerance (Cock and ElSharkawy 1988a,b; El-Sharkawy, 1993, 2004). The breeding objectives included: (i) characterisation of the core germplasm for tolerance to extended water shortages, either naturally or imposed, and to low-fertility soils; (ii) study of leaf photosynthetic potential in relation to productivity under various edaphoclimatic conditions; and (iii) identification of other plant traits that might be of use in breeding programmes. The multidisciplinary research approach adopted was pivotal in achieving these objectives. Selection of parental materials for tolerance to water stress and infertile soils has resulted in breeding improved germplasm adapted to both wet and water stress environments. A set of elite cassava germplasm with drought-tolerant attributes has been selected for detailed phenotypic analysis under water stress. The International Fund for Agricultural Development (IFAD, ROME) has funded a long-term project for selection in different semi-arid environments in Northeastern Brazil (where the greatest genetic diversity of cassava germplasm for adaptation to drought is found, see El-Sharkawy, 1993) and distribution of the elite germplasm throughout Africa (Fukuda and Saad, 2001). Farmers had participated in evaluating and selecting adapted germplasm that resulted in rapid acceptance and ,consequently, in the release of several improved clones (Saad et al., 2005).


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Responses of Field-Grown Cassava to Air Humidity and Its Implications for Breeding Strategies in Different Edaphoclimatic Zones and Ecosystems Cassava stomatal sensitivity to atmospheric humidity was observed at two locations in Colombia: in field-grown cassava in wet soils at the mid-altitude CIAT Headquarters station at Palmira, Valle Department, and at the low-altitude Carimagua ICA-CIAT station, Meta Department (Berg et al, 1986; Cock et al 1985; El-Sharkawy 1990). Over a range of cultivars representing the core cassava germplasm from different habitats and grown at a mid-altitude CIAT experimental station at Santander de Quilichao, Cauca Department, Colombia, significant differences in stomatal sensitivity to humidity were observed among cultivars (Figure 8, El-Sharkawy 2004). Furthermore, total biomass and storage root yield were greater in high humidity environments enhanced by misting that resulted in higher leaf photosynthesis, indicating that stomatal sensitivity to changes in VPD was translated at the canopy level (Figure 8, Cock et al 1985; El-Sharkawy and Cock 1986). Recent research on whole-plant water relations of field-grown cassava under prolonged natural water deficit in Ghana, West Africa, showed that both canopy conductance and transpiration declined with increasing VPD (Oguntunde 2005). These findings have important practical implications for cassava breeding and improvement for different ecosystems and edaphoclimatic zones. In wet/humid zones such as the Amazonian basin, equatorial west Africa, west Java in Indonesia, and in zones with short intermittent water deficits, less sensitive cultivars (with a lower stomatal density on the abaxial surface of hypostomatous leaves and/or with amphistomatous leaves with equal total conductances) should be bred for and selected in order to maximise productivity, since optimising WUE is not of importance in this situation (El-Sharkawy, 2004; El-Sharkawy and Cock, 1986)( more information on ontogenesis of leaves and the impact of stomatal density, size and pattern of distributions on both leaf sides, in terms of photosynthesis as well as the comparative adaptive advantages of amphistomatic versus hypostomatic characteristics can be found in: Gutschick, 1984; Mott et al, 1982; Parkhurst, 1978; Pospisilova and Solarova, 1980; Tichá, 1982). On the other hand, in subhumid/seasonally dry and semi-arid zones where there are prolonged periods of water deficit (more than three months), it is advantageous to breed and select for more sensitive cultivars in order to conserve the limited soil water and deplete it slowly, thus optimising WUE rather than maximising productivity, over a longer period during the growth cycle. In this case weed control must be practiced to avoid competition for water. Also, plant residual mulch, if available, would lead to better water conservation for crop use (Cadavid et al., 1998). Since new leaf formation is very restricted under prolonged drought (Connor and Cock, 1981; El-Sharkawy and Cock, 1987b; El-Sharkawy et al, 1992b; Porto, 1983), higher degrees of stomatal sensitivity should be combined with greater leaf retention, ie, longer leaf life and duration (El-Sharkawy, 1993, 2004). Recently, leaf retention was found to be positively correlated, over a wide range of cultivars and breeding lines, with productivity under naturally extended water deficits (Lenis, et al 2006). Moreover, it had been reported that leaves of plants subjected to prolonged imposed water stress (more than two months) in the field in subhumid zones had 40 percent of the net photosynthesis of well-watered plants and were capable of recovering completely after termination of the stress (CIAT, 1987, 1988, 1989,1990,1991, 1992,1993, 1994; El-Sharkawy, 1993). Selection for longer leaf lifespan is advantageous in saving dry matter already invested in leaf canopy formation (Chabot and

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Hicks, 1982), resulting in more assimilates being diverted toward storage roots, and leading to a higher HI and harvestable yield (Cock and El-Sharkawy, 1988a; El-Sharkawy, 1993). De Tafur et al (1997b) reported a wide variation in net leaf photosynthesis among rainfed cassava, as measured in the field during the driest months in a seasonally dry zone (Pn ranged from 27 to 31µmol CO2 m−2 s−1) and in a semi-arid zone (Pn ranged from 7 to 20 µmol CO2 m−2 s−1). Significant differences were observed among cultivars that could be exploited in breeding improved genotypes. In this case, host-plant tolerance/resistance for pests and diseases must be incorporated into cultivars targeted to seasonally dry and semi-arid zones in order to maintain functioning leaf canopy as much as possible over extended periods of time (Bellotti, 2002; Byrne et al, 1982; Calvert and Thresh, 2002; Hershey and Jennings, 1992; Hillocks and Wydra, 2002). There is also a wide genetic diversity in stomatal density within cassava germplasm, with a small percentage of the genotypes possessing amphistomatous leaves that may be used in breeding programmes. Thus, several accessions have been identified with a significant number of stomata on the upper surface, but these are less than 5 percent of more than 1,500 landraces and cultivars that were screened in the field. Screening was done using the transient porometer technique and microscopic observation on replicas of leaf surfaces made by spraying leaves with a collodion solution (El-Sharkawy et al, 1984a; 1985; Guzman, 1989). Both porometry (Kirkham 2005) and leaf surface replicas combined with microscopic observation are easy to handle in screening large amounts of breeding material in the field for stomatal characterisation, although the leaf replica method has some limitations in the case of hairy leaves and sunken stomata (North, 1956; Slávik, 1971). The leaf surface replica method was effective using young tissue-cultured seedlings (El-Sharkawy et al, 1984a). This may facilitate early screening of large populations. Zelitch (1962) described a similar technique using silicon rubber combined with a cellulose acetate solution to obtain stomatal impressions.

Screening Cassava Germplasm for Leaf Photosynthesis Having pointed out the importance of field research and the need to assess cassava potential photosynthesis under representative environments, we studied photosynthesis of recently matured upper canopy leaves in groups of cultivars, landraces and improved CIAT breeding material grown in three sites used by the cassava breeding programme for evaluation of genetic performance. The objective was to identify cultivars and lines with high photosynthetic potential in the field to be used as parental materials in crosses and breeding processes for improved productivity (El-Sharkawy, 1993), in combination with the other main breeding objectives of yield stability, broad-adaptation and tolerance/resistance to edaphoclimatic stresses, pests and diseases (Hershey and Jennings, 1992; Jennings and Iglesias, 2002). This objective was warranted, since our previous research in different locations in subhumid, seasonally dry and semi-arid environments showed a significant correlation, across a wide range of core germplasm and edaphoclimatic conditions, between upper canopy leaf photosynthesis – as measured in the field with portable infrared gas analysers – and total biomass and storage root yields (Figure 9; CIAT, 1987-1995; De Tafur, 2002; De Tafur et al, 1997b; El-Sharkawy, 2004; El-Sharkawy and Cock, 1990; El-Sharkawy et al, 1990, 1993; Pellet and El-Sharkawy, 1993a).


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Figure 9. Relationship between dry root yield and upper canopy leaf photosynthesis (A) and intercellular CO2 (B) for groups of cultivars grown in Santander de Quilichao (subhumid), Santo Tomas (seasonally dry) and Riohacha (semi-arid). Sources: El-Sharkawy,2006; De Tafur, 2002; De Tafur and El-Sharkawy, unpublished observations; De Tafur et al, 1997b; El-Sharkawy et al, 1993.

Tables 6, 7 and 8 present data on upper canopy leaf photosynthesis measured in the highaltitude cool climate (at Cajibio, Cauca Department, Colombia, elevation ca 1,800m, mean annual temperature ca 19°C) and the mid-altitude warm climate (at the CIAT Quilichao experiment station, Cauca Department, and at CIAT Headquarters, Palmira,Valle Department, with elevations about 965–1,000m, and a mean annual temperature of ca 24°C). Crops were grown under rain-fed conditions with minimum fertilizer applications. Measurements were

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made on several occasions, mainly during dry periods, and then averaged. Chambers enclosing whole or part of central leaf lobes were always directed toward the sun and at photon flux densities greater than 1,000µmol m−2 s−1 between 0900 and 1200h local time on four- to six-month-old plants, when the leaf canopy was nearly closing up (high leaf capacity source) and storage root bulking was at its highest rates (high root sink demand). In all locations, the average leaf photosynthesis varied significantly among screened cultivars and landraces, with rates greatly reduced at the high-altitude cool climate, confirming results and patterns observed with potted cassava (compare data of Tables 6, 7 , 8 and Figure 11). The accessions evaluated in the high-altitude cool climate were all local traditional cultivars or landraces from cool climate regions collected from several countries and included improved CIAT materials bred and selected for better adaptation to this climate. Compared to the overall mean photosynthetic rate (12.3µmol CO2 m−2 s−1), the few higher ranking materials with rates from 15.7–17.3µmol CO2 m−2 s−1 were four CIAT improved clones and a Peruvian cultivar (M Per 501) as shown in Table 6. These findings indicate the narrow genetic base for this ecosystem. They also confirm the relative effectiveness of the breeding strategy adopted by the cassava programme at CIAT for specific edaphoclimatic zones and ecosystems as discussed above, and point to the importance of including leaf photosynthesis as a selection criterion in parental materials for enhancing productivity (El-Sharkawy, 2004; El-Sharkawy and Cock, 1990; El-Sharkawy et al, 1990). Table 6. Leaf net photosynthesis (Pn; µmol CO2 m−2 s−1), stomatal conductance (gs; mmol m−2 s−1) and internal CO2 (Ci ; µmol CO2 mol−1) for some selected cassava clones with relatively high photosynthetic capacity. Plants were grown in 1992 and 1993 on a private farm at Cajibio, Cauca Department, Colombia (altitude 1,800m; cool subhumid climate). Measurements were made with portable infrared CO2 analysers on recently matured upper canopy leaves of 4–6-month-old plants. Note the higher Pn rates and the lower values of stomatal conductance and of internal CO2 in this group of clones, as compared with the means of the trial. This indicates the importance of non-stomatal (ie, biochemical/anatomical) factors in selection for enhanced photosynthetic capacity. There is a need to improve the genetic base for the cooler ecosystems in the high altitude tropics and in the subtropics where cassava is grown. Sources: Cassava Physiology Section database; CIAT, 1994 Selected clones

Pn

SM 1061-1

17.3

Stomatal conductance (gs) 196

SM 526-12

16.7

320

154

SM 1054-4

16.6

225

114

M Per 501

16.4

391

166

SM 1053-9

15.7

225

122

Mean

16.5

271

131

Mean of all accessions (n=107)

12.3

312

183

LSD 5%

1.3

32

14

Internal CO2 (Ci) 98


169

The enhanced photosynthesis in the few superior clones (Pn was 134% of overall mean of all accessions) could not be attributed to stomatal control, since their average stomatal conductance (with 271 mmol m−2 s−1) was significantly lower than the overall mean of all accessions (with 312 mmol m-2 s-1) (Table 6). However, the intercellular CO2 concentration was greatly reduced in these clones (Ci was 71% of the overall mean of all accessions), indicating possible control by non-stomatal factors such as leaf anatomy and biochemistry (eg, enzyme activity). Since the rate of leaf formation is much slower in a high-altitude cool climate but leaf life is much longer compared to warm climate conditions (Irikura et al, 1979), selection for enhanced photosynthesis and tolerance to low temperatures becomes even more important. Table 7. Leaf net photosynthesis (Pn) (µmol CO2 m−-2 s−1) in upper canopy leaves of the cassava core collection grown at Santander de Quilichao in 1993–1994. Measurements were made with a portable infrared gas analyser from 5–6 months after planting. Values are means of 7–11 measurements made during the dry period. These higher values of Pn obtained in a warm subhumid habitat can be compared with those obtained in a cool subhumid habitat shown in Table 6.The cultivar M Mal 48 from Malaysia had the highest Pn rate and the highest dry root yield in this trial. Sources: Cassava Physiology Section database; CIAT, 1994 Clone M Mal 48

Pn 27.6

Clone M Tai 1

Pn 24.4

M Bra 900

27.6

M Pan 51

24.3

M Bra 12

26.8

M Bra 383

24.2

M Bra 191

26.7

M Ind 33

24.1

M Mal 2

26.4

CM 849-1

23.6

HMC-1

26.0

M Col 1684

23.4

MCol 1468

25.8

M Mex 59

23.2

MCol 2061

25.4

M Ven 25

23.2

M Gua 44

25.4

M Bra 885

23.1

M Chn 1

25.3

M Cub 51

22.8

M Col 22

25.1

M Col 2215

22.3

M Arg 13

25.0

M Cub 74

22.3

M Ven 45A

24.8

M Per 205

22.0

M Col 1505

24.8

M Ptr 19

21.3

M Bra 110

24.6

M Ecu 82

21.0

LSD (5%)

4.8

In the mid-altitude warm climate sites, average photosynthesis rates were much greater than in the cool climate, particularly at the CIAT Headquarters location (Tables 7 and 8).The measurements were all made during the dry period. Thus, they were lower than the maximum rates observed in wet conditions (see Table 9). The majority of the materials evaluated at CIAT Headquarters were a collection of cultivars and landraces from Brazil, plus eight accessions from Argentina and one each from Colombia (HMC-1) and Bolivia (M Bol 1).

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The mean rate of photosynthesis was significantly greater in the few accessions from Argentina (26µmol CO2 m−2 s−1) compared to the germplasm from Brazil (Table 8).This was due to the high number of Brazilian accessions with rates lower than their origin overall mean of 22µ mol CO2 m−2 s−1. Nevertheless, there were several Brazilian accessions within the highest photosynthesis range, particularly M Br 12 (also see Figure 11B) and M Br 110, that could be used in crossing and breeding for warm climate ecosystems. Since the accessions from Argentina are, presumably, more adapted to subtropical ecosystems than the warm climate germplasm from tropical ecosystems and have some tolerance to low winter temperatures, they could be used in crossing and breeding for enhancing photosynthesis in high-altitude cool climate germplasm. In this group of clones, Pn was highly negatively correlated with intercellular CO2 (r2 =- 0.97, P < 0.0001; regression: intercellular CO2 = 315 - 7.83 Pn, Figure 10 ), indicating that differences in Pn were caused by non-stomatal factors, that is, anatomical and/or biochemical factors such as enzyme activity and leaf anatomy (El-Sharkawy et al. 1990, 2008; de Tafur et al. 1997b; El-Sharkawy 2006, 2010). The accessions screened at Quilichao were a mix of cultivars and landraces from Latin America, which were the majority, and some from Asia (Table 7). Again average photosynthesis rates varied widely among cultivars, with several high-ranking accessions from Brazil, Colombia and Malaysia. The highest ranking accession from Malaysia, M Mal 48, also had the highest dry root yield (15.6t ha−1) compared with the overall mean of the trial (10.6 tha−1). This clone had already been used in crossing and breeding at CIAT.

Figure 10. Relationships between Pn and intercellular CO2 concentration (Ci) in 53 cassava accessions (data of Table 8). Note the negative correlation between P n and Ci, which indicates that the association was caused by non-stomatal factors (i.e., biochemical and/or anatomical mesophyll traits) (SM de Tafur and MA El-Sharkawy 1995, unpublished).

Nevertheless, during rainy period maximum photosynthetic rates of several cultivars of field-grown cassava were more than 40 µmol CO2 m-2 s-1, with a mean Ci /Ca ratio of 0.42


171

(Table 9). These values are comparable with those observed in C4 species and much less than those obtained in C3 species. Table 8. Leaf net photosynthesis (Pn) in µmol CO2 m−2 s−1 of upper canopy leaves and intercellular CO2 (Ci) (µmol CO2 mol−1) for 53 accessions of the core collection grown at CIAT Headquarters in 1991–1992. Measurements were made with portable infrared CO2 analysers during dry periods in four-month-old plants. These higher Pn rates in warm subhumid habitat can be compared with rates obtained in a cool subhumid habitat as shown in Table 6. Note that few accessions from Argentina and Brazil have relatively high Pn rates as compared to the trial mean. (Cassava Physiology Section database; CIAT, 1992.) Accession

Pn

M Arg 11 M Bra 12 M Bra 110 HMC-1 M Arg 2 M Bra 132 M Bra 172 M Arg 13 M Bra 359 M Bra 71 M Arg 7 M Bra 85 M Arg 9 M Bra 190 M Bra 124 M Bra 403 M Bra 162 M Arg 5 M Bra 242 M Bra 299 M Bra 165 M Bra 243 M Bra 405 M Bra 73 M Bra 217 M Bra 404 M Bra 125 LSD 5%

32 30 30 29 28 28 27 27 27 26 26 26 26 26 26 25 25 25 24 24 24 23 23 23 23 23 23 6.2

Intercellular CO2 (Ci) 73 82 74 87 95 88 83 105 127 111 119 112 110 97 100 131 108 115 133 133 114 132 143 134 140 151 129 23

Accession

Pn

M Bra 258 M Bra 273 M Bra 77 M Bra 233 M Arg 12 M Bra 416 M Arg 15 M Bra 356 M Bra 191 M Bra 383 M Bol 1 M Bra 309 M Bra 453 M Bra 400 M Bra 329 M Bra 435 M Bra 337 M Bra 158 M Bra 325 M Bra 237 M Bra 355 M Bra 450 M Bra 328 M Bra 311 M Bra 315 M Bra 335

22 22 22 22 22 22 22 22 21 21 21 21 21 21 20 20 19 19 18 17 17 17 17 16 16 14

Intercellular CO2 (Ci) 143 128 145 144 146 151 139 156 145 162 147 148 159 169 159 173 177 137 169 173 195 196 182 188 191 187

172


Figure 11 A and B. Responses in cassava of net photosynthetic rate (P n) to leaf temperature for: (A) cv M Col 2059; habitat: cool humid; (B) cv M Bra 12; habitat; hot humid. ∆ = leaves developed in a cool climate; ● = leaves developed in a cool climate and then acclimated for one week in a warm climate; ๐ = newly developed leaves in a warm climate of the same plants. The apparent upward shift in optimum temperature from cool to warm-acclimated and warm climate leaves reflects the adaptation to warm climate in both cultivars. Also note: (i) the lack of light saturation (11C) in warm climate leaves as compared to cool and warm- acclimated leaves indicating the high capacity for CO2 fixation; (ii) the higher maximum photosynthetic rates in all sets of leaves of cv MBr.12 (11B)from hot-humid habitat compared to the cool-climate cv M Col 2059(9A). Sources: CIAT, 1992; El-Sharkawy et al, 1993; also see El-Sharkawy et al, 1992a.


173

Figure 11C. Respnses in cassava of net photosynthetic rate (P n) to irradiance (PAR) in cv M Col 2059. ∆ = leaves developed in a cool climate; ● = leaves developed in a cool climate and then acclimated for one week in a warm climate; ๐ = newly developed leaves in a warm climate.Sources: CIAT, 1992; El-Sharkawy et al, 1993; also see El-Sharkawy et al, 1992a.

Table 9. Leaf net photosynthesis, Pn, of field-grown cassava at Santander de Quilichao, Cauca Department (warm subhumid) during the 1990–991 season. Maximum photosynthetic rates were obtained during wet periods and high air humidity. Measurements were made on upper canopy leaves using portable infrared CO2 analysers. The Ci/Ca ratio (intercellular CO2 divided by atmospheric CO2). is commonly used to differentiate plant species based on their photosynthetic capacities; ie, the lower the ratio the higher the capacity. Note the Ci/Ca values here which are comparable to those in C4 species and much lower than those in C3 species, indicating the high photosynthetic capacity of cassava as expressed in near optimum environments. In this group of cultivars, average seasonal Pn was correlated with final root yield. Sources: Cassava Physiology Section database; El-Sharkawy et al, 1992a, 1993

CG 996-6

Maximum net photosynthesis µmol CO2 m-2 s-1 (n = 6) 49.7

0.37

Seasonal average net photosynthesis µmol CO2 m-2 s-1 (n = 30) 33.8

M Bra 191

47.4

0.37

35.5

CM 4864-1

45.1

0.39

34.0

CM 4145-4

43.9

0.40

31.7

CM 3456-3

43.7

0.43

31.9

Cultivar

Ci/Ca (n = 6)

174

Mabrouk A. El-Sharkawy, Sara M. de Tafur and Yamel Lopez Table 9. (Continued)

Cultivar

Maximum net photosynthesisµmol CO2 m-2 s-1 (n = 6)

Ci/Ca (n = 6)

Seasonal average net photosynthesisµmol CO2 m-2 s-1 (n = 30)

CM 507-37

43.7

0.38

28.7

CM 4716-1

43.6

0.42

31.8

M Col 1684

43.0

0.42

30.9

CM 4575-1

42.8

0.39

33.2

CM 4617-1

42.8

0.46

31.4

CM 523-7

42.3

0.45

30.1

M Col 1468

42.3

0.44

30.3

CM 4701-1

42.2

0.45

30.9

CM 4711-2

41.3

0.45

30.9

CG 927-12

39.3

0.43

26.2

Mean of all cultivarsss

43.5

0.42

31.4

LSD (0.05)

1.70

0.08

1.80

Evaluation of Cassava Core Germplasm for Leaf Area Duration (Seasonal Average Leaf Area Index) and Productivity in a Mid-Altitude Warm Climate To complement the joint physiology and breeding efforts in characterising cassava core germplasm and in the identification of useful yield-determinant traits, a field trial was conducted at the CIAT mid-altitude warm climate experimental station at Quilichao where 30 clones were evaluated for leaf duration over the growth cycle by measuring the seasonal average LAI using a leaf canopy analyser. Data on yield, shoot and total biomass, seasonal average LAI and root dry matter content are given in Table 10 (CIAT, 1995). Wide variations among clones were found in standing shoot biomass (top, excluding fallen leaves) and total biomass, yield, dry matter content of roots and seasonal LAI. It is noteworthy that several accessions from Brazil were among the highest ranked in terms of yield, total biomass and dry matter content in storage roots, thus highlighting the importance of the Brazilian germplasm. Breeding at CIAT, while diversifying the genetic base, has incorporated many of these accessions for their useful traits. Outstanding among these accessions is the clone M Br 12, with its high leaf photosynthesis in cassava grown both outdoors in pots or in the field in a mid-altitude warm climate (see Figure 11B and Table 8), and its high yield coupled with resistance to mites (Byrne et al, 1982). Other accessions of Brazilian origin, M Br 383 and M Br 191, that ranked high in this group of clones, were also reported to be among the highest ranked clones (fourth and fifth, respectively, among 33 evaluated) for tolerance to soils low in phosphorus (CIAT, 1990; ElSharkawy, 2004).


175

Table 10. Seasonal average LAI, dry root yield, top and total biomass, and root dry matter content of 30 clones from the cassava core collection. Plants were grown at Santander de Quilichao in 1994–1995. Leaf area duration, as estimated by seasonal average LAI, was significantly correlated with root yield (see Figure 12), indicating the importance of this trait in breeding and selection for improved cultivars. Sources: Breeding and Physiology Sections database, CIAT,1996

M Bra 383 M Bra 12

Seasonal average LAI m2 m-2 1.3 1.0

Dry top biomass t ha-1 6.0 5.3

Dry root 15.5 15.3

Total biomass 21.5 20.6

Root dry matter % 41.7 38.2

M Pan 51

1.8

10.2

14.7

23.9

40.8

CM 849-1

1.4

6.3

14.1

20.4

41.7

M Bra 191

1.5

5.7

14.0

19.7

41.3

M Mal 48 M Bra 885

1.9 1.6

4.2 5.4

13.9 13.8

18.1 19.2

41.0 41.6

M Ven 25

1.1

4.9

12.6

17.5

40.0

HMC-1

1.6

4.8

12.2

17.0

38.5

M Ind 33

1.6

5.9

11.6

17.5

36.6

M Bra 100

1.9

6.4

11.3

17.7

40.6

M Gua 44 M Cub 74

1.2 0.9

4.6 3.9

11.2 10.8

15.8 14.7

38.9 39.8

M Tai 1

1.2

5.6

10.6

16.2

37.5

M Mex 59

1.6

6.8

10.3

17.1

40.7

N Arg 13

0.9

2.4

9.3

11.7

39.1

M Mal 2

1.8

6.9

8.4

15.3

36.2

M CHN 1 M Col 22

1.0 1.4

1.9 1.7

8.0 7.8

9.9 9.5

36.2 37.8

M Ven 45A

1.0

5.1

7.7

12.8

37.4

M Col 1684

0.8

2.2

7.0

9.2

34.6

M Ecu 82

0.8

3.4

6.7

10.1

38.6

M Ptr 19

1.1

4.0

6.1

10.1

40.0

M Col 2215 M Col 1468

0.7 0.9

2.4 1.9

5.9 5.6

8.3 7.5

40.3 36.7

M Col 2061

0.9

3.5

5.3

8.8

30.7

M Per 205

1.0

3.8

5.3

9.1

38.7

M Col 1505

0.7

2.1

5.1

7.2

40.6

M Cub 51

0.7

3.1

4.9

8.0

39.9

M Bra 900 Mean of all clones

0.9 1.2

1.3 4.4

4.7 9.7

6.0 14.0

31.4 39.6

LSD 5%

0.5

2.0

3.5

4.6

3.6

Clones

In this group of accessions, standing shoot biomass correlated with root yield (r = 0.70; p < 0.001), confirming previous findings that suggest using this trait as a proxy for leaf area formation and duration when evaluating large breeding populations (CIAT, 1990; ElSharkawy, 2004; El-Sharkawy et al, 1990).

176

Mabrouk A. El-Sharkawy, Sara M. de Tafur and Yamel Lopez 18

-1

Dry root yield (t ha )

16 14 12 10 8 y=2.34 + 6.07x r=0.65 (p=0.001)

6 4 0

1

2 LAI

Figure 12. The relationship between final dry root yield and seasonal average LAI in 30 clones from the core cassava germplasm collection at CIAT. Crops were grown at the CIAT Quilichao station with a mid-altitude warm climate. Leaf area was determined throughout the growth period using a leaf canopy analyser. The significant correlation indicates the importance of leaf area duration for yield formation. Due to the small leaf area canopy in the first 1–3 months and in the final 8–12 months, the seasonal average canopy area still limits yield and indicates the need to breed and select for higher and more sustainable canopy during most of the growth cycle, combined with enhanced leaf photosynthesis, high HI and a strong root sink (a higher storage root number per plant). It is noteworthy that cv M Br 12 had a high yield with an LAI lower than the overall average of the trial, indicating its high leaf photosynthetic potential (also see Figure 11B). Source: CIAT, 1995.

Also, as shown in Figure 12, dry root yield correlated with seasonal LAI in this group of clones (r = 0.65, p < 0.001). This further substantiates earlier reports (Pellet and ElSharkawy, 1993a) and supports the concept of breeding for longer leaf life and optimum leaf area duration, for maximising productivity under favorable conditions as well as for sustainable yields in stressful environments (Cock and El-Sharkawy, 1988a,b; El-Sharkawy, 1993, 2004; El-Sharkawy and Cock, 1987b; El-Sharkawy et al, 1992b; Lenis et al, 2006). In conclusion, the use of leaf photosynthesis as a selection criterion in cassava improvement programs might be difficult to handle when evaluating large breeding populations. Despite this, it should be included at least in the processes to evaluate and select parental materials, in combination with other important yield-related traits, particularly relatively high HI (> 0.5; Kawano, 1990, 2003), large root sink using root number per plant as a criterion (Cock et al, 1979; Pellet and El-Sharkawy, 1993a,1994), and longer leaf life, ie, greater leaf retention and duration over the growth cycle (Cock and El-Sharkawy, 1988a,b; El-Sharkawy, 1993, 2004; El-Sharkawy and Cock, 1987b; El-Sharkawy et al, 1992b; Lenis et al, 2006). Recent advances in the area of molecular biology and in the development of more precise techniques and equipment,will further enhance and speed up elucidation of the fundamental mechanisms underlying photosynthetic potential and associated beneficial traits and their controlling genes (Daniell et al., 2008; Raines 2006,2011).


177

Evaluation of Germplasm under Mid-Season Water Stress in a Field Drainage Lysimeter Sixteen cultivars from the cassava core germplasm collection were evaluated over a number of years in a field drainage lysimeter under three months of water stress initiated 90– 100 days after planting (ie, mid-season stress). Figure 13 illustrates dry root yield accumulation patterns for four representative accessions as affected by stress during the growth cycle.

Months after planting

Figure 13. Dry root yield in a group of clones as affected by a three-month water stress period initiated 90 days after planting (ie, mid-season stress). The yield was significantly lower at the end of stress, but recovered rapidly with watering and the final yields approached those of the controls. There were differences among cultivars with CM 489-1 having the highest yield in both water regimes (also see Table 11). It is noteworthy that CM 489-1 had high leaf photosynthesis with the least reduction under stress (see Figures 3 and 14) and high PEPC activities in leaves of fieldgrown crops under extended water stress. Sources: CIAT, 1992; El-Sharkawy, 2004.

Water stress significantly reduced yield in all accessions at the end of stress, as well as reducing shoot biomass (CIAT, 1991, 1992; El-Sharkawy et al, 1992b). However, after recovery from stress, final yields were equal to those of well-watered plants in some accessions, while reduced in others (see Table 11; El-Sharkawy, 1993). There were also significant differences among cultivars in final root yield, with the hybrid CM 489-1 having the highest yield under both stress and non-stress conditions (18t ha1 and19t ha-1 respectively of oven dry roots). It is noteworthy to report here that CM 489-1

178


also showed high PEPC enzyme activity in leaf extracts under extended field water shortages. This correlated with leaf photosynthesis across a group of cultivars (El-Sharkawy, 2004), high values of yield, leaf photosynthesis, NUE in terms of root production, radiation-use efficiency (RUE) in terms of total biomass production, and a large number of harvestable storage roots per plant across a range of phosphorus fertilizer levels in acidic soils in a subhumid warmclimate (Pellet and El-Sharkawy 1993a,b, 1994, 1997). Across accessions, reductions were much greater in shoot biomass (28 percent) compared to roots (9 percent), with about 6 percent increases in HI, indicating the potential of cassava to tolerate prolonged mid-season stress in subhumid zones. It also indicates its ability to recover and compensate for possible losses in productivity, giving it an advantage over other staple food crops (CIAT, 1991, 1992; El-Sharkawy and Cock 1987b; El-Sharkawy et al 1992b,). There is genetic variability in tolerance to stress that should be exploited in breeding and improvement of cassava germplasm for dry environments (CIAT, 1991, 1995; El-Sharkawy, 1993; Hershey and Jennings, 1992). Table 11. Root yield and top biomass, and hydrogen cyanide (HCN) content in roots at final harvest (11 months) as affected by three months of mid-season water stress commencing 90–100 days after planting. Data are the average of the 1987–1989 seasons at Santander de Quilichao. Note the increase in HCN content due to stress and the differences among cultivars. Clones with lower HCN under stress are good genetic sources for breeding and selection for suitable materials for fresh consumption by humans, particularly in dry and semi-arid zones. Sources: Cassava Physiology Section database; CIAT, 1991; El-Sharkawy, 1993 Unstressed Clone

CM 489-1 CM 922-2 CM 1335-4 CM 2136-2 Average % change due to stress

Biomass (dry t ha−1) Roots

Tops

Total HCN (mg kg−1 dry root)

19.1 14.8 18.1 19.3 17.8

7.2 7.6 7.8 12.4 8.8

214 142 107 166 157

Stressed Biomass (dry t ha−1) Roots

Tops

Total HCN (mg kg−1 dry root)

18.0 15.0 16.5 15.5 16.2 −9

7.1 5.9 5.1 7.3 6.4 −28

401 190 123 338 263 +68

Most cassava cultivars show increases in hydrogen cyanide (HCN) content – as an indicator of cyanogenic potential – in their storage roots when exposed to extended water deficits, thereby becoming less suitable for human consumption if not properly processed to remove most, if not all, HCN (Dufour, 1988; Essers, 1995; Rosling,1994). Some crop management practices, such as applying moderate amounts of NPK fertilizers and/or plant residues as mulch to infertile sandy soils in zones with long dry periods (Cadavid, et al 1998), or applying potassium to clayey acidic soils low in potassium in subhumid zones (ElSharkawy and Cadavid, 2000), will greatly reduce HCN in cassava roots. However, selection


179

for low HCN cultivars remains a main objective in most breeding programmes, particularly for germplasm targeted to stressful environments (El-Sharkawy, 1993). In this case, the identification of less sensitive and low HCN genotypes as shown here (Table 11) offers a good genetic source for breeding sweet cultivars. Nassar (1986, 2000) has also reported some wild species with low HCN and high protein in their storage roots. Such wild species should be used as genetic sources in future research. Acyanogenic (cynogen-free) line of cassava was recently genetically engineered (Siritunga and Sayre, 2003) and is currently under field evaluations. Also, transgenic plants of cultivar MCol 22 with a 92% reduction in cyanogenic glucoside content in storage roots and with acyanogenic leaves were successfully produced (Jørgensen et al., 2005), which might be used either directly (after agronomic evaluations in the field) or in crosses with high yielding and drought-tolerant cultivars such as MCol 1468 (also named CMC 40) (Figure, 15) and CM 489-1(Table 11). The cultivar MCol 22 was previously reported to be less drought tolerant (Connor et al., 1981; El-Sharkawy and Cock, 1987b). Despite these reports, others (Pereira, 1977; Poulton, 1990) argue that high levels of HCN in plants may play a role as a defensive mechanism in protecting crops against predators and herbivores or serves as a source of stored nitrogen, particularly in seeds. However, a putative defensive role ascribed to HCN has not been reflected in the incidence of cassava pests and diseases (Brekelbaum et al., 1978). For example, field observations in Northeastern Brazil and northern Colombia, where cassava crops experience several months of water shortage, showed higher mite infestations under water stress when leaf HCN is normally elevated in most cultivars, compared to lesser degrees of infestation in unstressed crops (El-Sharkawy, unpublished observations). Similarly, thrips was found to feed on cassava regardless of leaf HCN levels (van Schoonhoven, 1978). Other pests with different feeding habits, whether on shoot or on root, may present different responses. Work at CIAT (Bellotti, 2002; Bellotti and Arias, 1993; Bellotti and Riis, 1994; Bellotti et al, 1988) showed that the root-burrowing bug, Cyrtomenus bergi, preferred feeding on cassava roots low in HCN as compared to bitter cassava, particularly in wet soils, although several sweet cultivars were noted as having potential resistance/tolerance to the bug (Riis, 1997). One mechanism that may reduce attack or deter the bug from feeding on sweet cassava is a high HCN content in the storage root peel compared to the parenchyma tissue (Riis, 1997). Since the feeding habit of the bug’s first two nymphal instars with short stylets is confined mainly to the root peel (Riis, 1990; Riis et al, 1995), selection for sweet cultivars having high HCN in thicker root peel might be advantageous. Cultural practices such as intercropping cassava with the sunn-hemp Crotalaria sp, which possesses natural insecticidal substances, were found to effectively reduce bug attack and damage to cassava roots, although cassava yield was reduced due to competition and land occupation by Crotalaria (Bellotti et al. 1988; Bellotti 2002). In a recent study conducted in the northwestern Amazonian basin in Brazil with the native Tukanoan Indians, whose subsistence depends more on cultivating bitter cultivars high in HCN, no consistent relationship was demonstrated between the social preferences of cultivating bitter cultivars versus sweet ones and resistance to predators, particularly pests and diseases (Wilson, 2003).

180


Figure 14. Leaf photosynthesis in the upper canopy as affected by mid-season water stress. Measurements were made during the dry period using portable infrared gas analysers. Values are means of five to eight leaves from different plants. Note the progressive decreases in photosynthesis over time due to water stress and the recovery after termination of stress in old leaves, and the consistently higher rates in newly developed leaves in previously stressed crops as compared to the controls. There were apparent cultivar differences. Note: the least reductions in P n of CM 489-1, as compared with other cultivars. Source: CIAT, 1991.

Although dependence on bitter cultivars may be due to their predominance – sweet cultivars are less abundant – rather than to an inherent adaptive advantage, Wilson and Dufour (2002) reported that the higher yield often observed in bitter cultivars is the likely


181

criterion for the natives of the Amazonian basin to choose high HCN cassava. However, to our knowledge there is no conclusive evidence based on sound research of an inherent superior potential productivity in bitter cultivars compared to sweet ones. Data in Table 10 and those reported in 14 other cultivars tested for five consecutive growth cycles under different rates of application of potassium fertilizer in acidic clayey soils in the subhumid zone in Colombia (El-Sharkawy and Cadavid, 2000), indicated no consistent relationship between productivity and root HCN content. Moreover, several of the clones tested such as HMC-1, HMC-2, M Cub 74, M Pan 70, M Col 1505, CM 91-3,CM 523-7,CMC 40, CM 1585-13, as well as those shown in Table 10, have high yields and moderate to low HCN levels in the root parenchyma. Most of these clones are improved materials, thus indicating the compatibility of selection and breeding for both high yield and low HCN. Therefore, more research is warranted to uncover other possible reasons behind the choice of bitter cassava in the Amazonian region and elsewhere.

Evaluation of Germplasm for Tolerance to Prolonged Early Water Stress, Mid-Season Stress and Terminal Stress Three-year field trials were conducted at the CIAT Quilichao experimental station to study the effects of prolonged water stress imposed at different growth stages on cassava productivity, nutrient uptake and NUE, leaf photosynthesis and patterns of water uptake (Caicedo, 1993; CIAT, 1992, 1993; El-Sharkawy and Cadavid, 2002; El-Sharkawy et al 1998b). Figure 15 illustrates the dynamics of dry matter accumulation in storage roots over the growth cycle of four contrasting clones. Under early stress, initiated at two months after planting and terminated at six months, root yield at six months was significantly smaller than the control for all clones in both growth cycles. The same trends were observed in shoot biomass but with greater reductions than observed in roots (Caicedo, 1993; CIAT, 1992,1993; El-Sharkawy and Cadavid, 2002; El-Sharkawy et al, 1998b). Under mid-season stress, initiated four months after planting and terminated at eight months, yield at eight months was also significantly lower compared to the well-watered control in both cycles except for CMC 40 (or MCol 1468, its name in Colombia). Reductions in shoot biomass were less pronounced compared to those under early stress (Caicedo, 1993; El-Sharkawy and Cadavid, 2002; El-Sharkawy et al, 1998b). Leaf area, as determined from periodical harvests, was also significantly smaller in both early and mid-season stress compared to the controls, resulting in significantly reduced canopy light interception (Figure 16a, CIAT, 1992; El-Sharkawy and Cadavid, 2002). After the crop was allowed to recover, new leaf area formed rapidly, with values in early and mid-season stressed crops similar to or greater than controls, thereby resulting in greater light interception (Figure 16a). In the early stressed crops, increases in shoot biomass were less and remained lower than other water regimes, indicating adverse effects of the early stress on shoot biomass recovery (Caicedo, 1993; El-Sharkawy and Cadavid, 2002; ElSharkawy et al, 1998b).

182


Figure 15. Storage root dry matter yield (ton ha-1) in response to prolonged water stress imposed at different stages of growth: 2–6early, 4–8 mid-season, 6–12 terminal, months after planting in four cassava cultivars. Crop cycles: a: first season(left panels), b: second season (right panels). ■ control, ○ early stress,  midseason stress, ▲ terminal stress. Note the reduction in yields during stress and the recovery at final harvest in early and midseason stress. There were cultivar differences with cv. CM 40 (M Col 1468) having higher yield under stress (El-Sharkawy and Cadavid 2002).


Figure 16. Photon interception (x102 %) (A–C) and net photosynthetic rate, Pn(D–F) of cassava crops (four cultivars) as affected by early (A), midseason (B), and terminal (C) water stress. ● stress, ○ control, ↓ stress start, ↑ recovery, DAP = days after planting. Note in A the large reduction in photon interception in early stress and the recovery in both early and midseason stresses that exceeded the control. Greater reduction was found in PN during terminal stress as compared to early and midseason stress treatments (CIAT 1992).

183

184


Under terminal stress initiated at 6 months after planting and remaining until final harvests at12 months, with the exception of CMC 40 whose yield was higher under stress, final root yields at 12 months were less than the control, with the largest reduction in CM 523-7 (or ICA Catumare, its name in Colombia). Genotype by water regime interaction was significant (p < 0.01), indicating the soundness of the strategy to breed and select for specific edaphoclimatic zones. Across clones in both years, final yields were not significantly different among water regimes (Table 12), indicating the capacity of cassava to tolerate extended water deficits in subhumid and seasonally-dry warm climates with bimodal precipitation patterns. It is noteworthy that clone CMC 40 had the highest yield and shoot biomass under terminal stress, with the smallest leaf area as compared to CM 523-7. This result could be attributed mainly to high leaf photosynthesis observed in the field under diverse environments (El-Sharkawy et al, 1990,1992a; Pellet and El-Sharkawy,1993a). Moreover, upper canopy leaves of CMC 40 showed greater activities of both the C3 and C4 main enzymes compared to CM 523-7. Values in µmol mg−1 chlorophyll min−1 were 8.2 for Rubisco and 3.1 for PEPC in CMC 40 versus 3.6 and 1.6 in CM 523-7 (Table 13, CIAT, 1992; López et al, 1993). These findings point to the importance of selecting and breeding for high leaf photosynthesis, particularly under stress. Variations among clones in leaf photosynthesis, as measured in the field, were observed (see Figure 16 D-F). Table 12. Effect of prolonged water stress imposed at different growth stages on storage root yield and shoot biomass at 12 months after planting, and on mean LAI over the growth cycle for four cassava cultivars grown at Santander de Quilichao, in the 1990/91, 1991/92 and 1992/93 seasons. Data are for the second and third seasons. Note the highest root yield of cultivar CMC 40 (or MCol 1468) under prolonged terminal water stress and the highest activities of PEPC and Rubisco in the same cultivar as in Table 13. *NS = Not significant at 5%. Sources; Cassava Physiology Section database; Caicedo, 1993; El-Sharkawy and Cadavid, 2002; El-Sharkawy et al, 1998b

Terminal stress

5.3

2.3

1.3

1.8

2.3

CM 523-7

13.8

12.8

12.1

9.7

5.3

4.2

5.2

4.5

2.3

1.4

1.3

2.0

CMC 40

10.0

10.4

12.1

14.6

7.8

3.9

5.7

6.7

1.7

1.1

1.1

1.7

M Col 1684 Average

13.6

10.3

12.5

11.5

5.0

2.2

4.0

4.3

1.8

1.1

1.3

1.8

12.9

11.2

12.9

11.7

6.0

3.3

5.1

5.2

2.0

1.2

1.4

2.0

Control

Early stress

5.6

Terminal stress

2.7

Early stress

6.0

Control

11.1

Terminal stress

11.3

Early stress 11.1

Control

Mid-season stress

Treatment

14.0

Root yield (t ha-1 dry wt)

Mid-season stress

Treatment

CM 507-37

Cultivar

Mid-season stress

Treatment

Shoot biomass (t ha-1 dry wt)

Mean LAI

LSD 5% Treatment

NS*

0.8

0.3

Treatment x Cultivar

2.7

1.4

0.5


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Table 13. Activities of some photosynthetic enzymes in leaf extracts of field-grown cassava at Santander de Quilichao in 1992. Values are means ± SD. Note the wide range of genetic variations in enzymes activities that could be used in selection and breeding for enhanced photosynthesis and hence productivity. It is noteworthy that the cultivar MCol 1486 (or CMC 40) had the highest activities of both Rubisco and PEPC and the highest root yield under prolonged terminal water stress as shown in Table 12. Sources: Cassava Physiology Section database; CIAT, 1992; Lopez et al, 1993 Activities (µmol mg chl-1 min-1) PEPC

Rubisco

NAD-ME

NADP-ME

PEPC/ Rubisco

CM 523 – 7

1.57 ± 0.10

3.62 ±0.62

1.84 ± 0.1

0.41 ± 0.04

0.43

CM 507-37

1.91 ± 0.10

6.84 ± 0.66

1.37 ± 0.4

0.46 ± 0.18

0.28

M Col 1684

2.90 ± 0.19

6.96 ± 1.18

2.05 ± 0.6

0.36 ± 0.03

0.42

M Col CMC 40

3.07 ± 0.27

8.16 ± 0.71

1.84 ± 0.6

0.24 ± 0.05

0.38

Clone

Water Uptake and Water Use Efficiency Patterns of water uptake across clones during extended water stress imposed at different stages of growth are shown in Figure 17. In all stress treatments, cassava withdrew more water from the deep soil layer (two meters in depth), after the upper layer was almost depleted. Moreover, the water uptake from this deep layer increased as stress progressed, particularly in the terminal stress treatment, indicating the deep rooting behaviour as previously reported (CIAT, 1991, 1992, 1993, 1994; Cadavid et al, 1998; Connor et al, 1981; De Tafur et al, 1997a; El-Sharkawy and Cock, 1987b; El-Sharkawy et al 1992b). It is noteworthy that the available soil water by volume in this soil ranges from 8 to 12 percent (see Connor et al, 1981; El-Sharkawy et al, 1992b).

Figure 17. Patterns of water uptake by cassava during extended water deficit at Santander de Quilichao. Data are averages of four cultivars. Note the greater water extraction from deeper soil layers that increased as water stress progressed over time, particularly in terminally stressed crops (El-Sharkawy et al. 1992b, CIAT 1993, De Tafur et al. 1997a, El-Sharkawy, 2006, 2007, 2010).

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Hence, the water uptake under any of these prolonged stress treatments would probably not exceed about 200mm (the total available water in two meters depth of soil), showing cassava’s capacity to conserve and deplete deep soil water slowly over an extended period of stress. In two cultivars (MCol 1684 and its hybrid CM 507-37) subjected to a three month midseason water shortage, El-Sharkawy et al (1992b) reported that total water uptake, as estimated from periodical sampling of soil cores at two meters depth during 35 and 96 days of stress was around 100 and 160mm respectively. The latter value is equivalent to 7075 percent of the plant-available water at this depth, and much less than the rate of pan evaporation of about 4.4mm day−1 at the site of the trials. In recent studies, using the sap flow method, Oguntunde (2005) estimated daily cassava canopy transpiration under prolonged natural stress in Ghana, at ca 0.8–1.2mm, which is equivalent to 24 percent of the potential evapotranspiration. El-Sharkawy et al (1992b) found that estimated crop WUE values during the 43-day period of peak canopy growth between 117 and 160 days after planting, were around 4.4– 4.8g kg−1 of water in the stressed crops as compared to 3.7–4.9g kg−1 in non-stressed crops for MCol 1684 and its hybrid CM 507-37, respectively (values being based on total ovendried biomass). Because cassava has a long growth cycle and a low LAI during a significant portion of its growing season, seasonal crop WUE is reduced. Nevertheless, estimated cassava seasonal crop WUE at about 2.9g total dry biomass per kg of water, in field lysimeter-grown crops compares favourably with values found with the C4 grain sorghum at ca 3.1g kg−1, and much greater than those in the C3 field bean at ca 1.7g kg−1 (El-Sharkawy, 2004; El-Sharkawy and Cock, 1986). Due to a higher HI in cassava (0.6–0.7), WUE in terms of economic yield would be even greater than in both sorghum and field bean, with their lower HI values. Moreover, in a large yield field-trial with 16 improved cassava cultivars grown in a seasonall dry zone in the Patia Valley, Cauca Department, Colombia, with less than 1,000mm of precipitation in 10 months, average harvestable total dry biomass, excluding fallen leaves and fine roots, was greater than 30t ha−1 (El-Sharkawy et al, 1990). In this trial, the mean oven-dried root yield among cultivars ranged from 15 to 27.4 t ha−1, with an overall mean of 20 t ha-1. If it is assumed that all rainfall was effectively used by these crops and lost via plant transpiration and evaporation from exposed soils only, ie evapotranspiration(although 60 percent of the rainfall occurred in the sixth and seventh months during the growth cycle, that may imply some water runoff and deep percolation into the clayey soil), an evapotranspiration ratio (water loss per unit total dry matter produced) of about 270–300 is then obtained. Such a ratio is comparable to values (excluding soil evaporation) observed in tropical C4 crop species such as maize, millet, sorghum and sudangrass, and much smaller than values obtained in C3 species (Stanhill, 1986). The actual evapotranspiration ratio of cassava may have been even lower than the estimated values since fallen leaves my account for 2-6 t ha−1 (El-Sharkawy and Cock 1987b; El-Sharkawy et al 1992b; Pellet and ElSharkawy, 1997). In terms of economic yield, an estimated evapotranspiration ratio of about 500 in this trial is much less than values in grain sorghum (868), proso millet (567) and maize (1,405) obtained in the Briggs and Shantz trials (see Stanhill 1986). If adjusted for moisture content in the grains of these cereals, the values would be much greater than in cassava. These findings indicate the potentially high WUE of cassava, and further strengthen its comparative advantage in water-limited zones.


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However, under non-limiting rainfall or with irrigation, cassava would be more efficient in terms of annual calories produced per unit land area and water consumption than most tropical C4 grain crops (El-Sharkawy, 1993, 2004). It has been predicted (El-Sharkawy, 2005) that productivity of cassava would probably be enhanced further due to rises in atmospheric CO2 and temperature (ie, the known global climate change phenomena), that should result in a much greater WUE. This prediction is substantiated by recent findings (Fernández et al, 2002) that leaf photosynthesis and root yield of field-grown cassava under elevated CO2 in the tropics (at 680cm3 m−3 within open-top chambers) were higher than values in ambient CO2grown plants, indicating the absence of the photosynthetic down-regulation often observed in some other species (see Kramer, 1981; Mooney, et al1991; Pettersson and McDonald, 1994; Webber et al, 1994; Woodrow, 1994). The absence of such down-regulation was apparently associated with greater carboxylation efficiency of the C3 Rubisco, despite reductions in soluble protein and nitrogen content in leaves, indicating higher nitrogen use efficiency in terms of CO2 uptake. El-Sharkawy (2004) reported significant positive correlations between dry root yield and photosynthetic nitrogen use efficiency – expressed as CO2 uptake per unit of total leaf nitrogen as measured in normal air and high solar irradiance in upper canopy leaves – across a wide range of field-grown cassava cultivars without CO2 enrichment. Furthermore, the soluble sugars and starch content in leaves of elevated CO2-grown plants were reduced, suggesting higher demands by strong sinks for assimilates, as indicated by the concomitant increases in both shoot biomass and storage root yield (Fernández et al, 2002). Also, Ziska et al (1991) reported a 56 percent increase in leaf photosynthesis of cassava plants grown for long periods in elevated CO2 under controlled conditions (at 300cm3 m−3 above ambient CO2) indicating the absence of down-regulation. These photosynthetic attributes combined with a high optimum temperature for leaf photosynthesis (see Figure 11 A, B) and elevated activities of the C4 PEPC in cassava leaves (see Tables 3, 4,5 and 13) may confer an adaptive advantage upon cassava growth and productivity in the case of global warming. In contrast to the above findings, Gleadow et al. (2009) reported lower leaf photosynthetic rates in plants grown at higher than ambient CO2, particularly in those grown at the highest level. They also found significant reductions in shoot and storage root biomass. However, we point out that these authors studied potted cassava grown under greenhouse conditions at different levels of CO2, that is, at 360, 550, and 710 ppm CO2. Obviously, these findings indicated a feedback inhibition due to the rooting systems being confined by the growth media used. Cassava is a tropical shrub (Figure 1) that requires large volumes of soil for the development of its storage roots. When grown under inappropriate conditions, it will not express either its leaf photosynthetic capacity or its potential productivity. Thus, as warned earlier in the chapter, such findings are invalid and any resulting conclusions must be questioned. In contrast, field research conducted in CIAT’s sunny, hot, humid, and tropical environment, demonstrated cassava’s high photosynthetic capacity and productivity. Cassava’s deep rooting characteristics, combined with partial stomatal closure in response to both soil and atmospheric water deficits, as well as a reduced leaf canopy and light interception while maintaining reasonable leaf photosynthesis, are of a paramount importance when the crop has to endure several months of prolonged drought in seasonally dry tropical ecosystems, where excess rains often recharge deeper soil layers. This pattern of conserved water use results in optimising seasonal crop WUE (El-Sharkawy, 2004; ElSharkawy and Cock, 1986). Boyer (1996) reviewed and discussed advances in drought tolerance in field crops and the possible mechanisms underlying enhanced crop WUE. The

188


deep rooting characteristics accounted for a major part of differences in drought tolerance among species. These inherent mechanisms may partially explain why stressed cassava is still capable of producing reasonably well, as compared to other cereal and grain legume food crops, and further strengthen the strategy of expanding cultivation of cassava in droughtprone areas where severe food shortages might occur (De Tafur et al, 1997a; El-Sharkawy, 1993, 2004; Hershey and Jennings, 1992).

SELECTION FOR SHORT-STEMMED CULTIVARS MORE EFFICIENT IN NUTRIENT USE AND IN LEAF PHOTOSYNTHESIS Research under prolonged water deficits in the field (El-Sharkawy et al., 1998b) showed that early and midseason water deficits increased nutrient use efficiency, in terms of root yield, due to more reduction in aerial parts (stems and leaves) as compared to storage roots, and hence, less total plant nutrient uptake. This observation had led to search for genotypic variations in productivity and nutrient use efficiency as affected by plant type. Among two groups of tall and short-stemmed cultivars, aerial parts were significantly lower in short-stemmed cassava while root yield was approaching values in tall cassava (Table 14). Seasonal average leaf area index (LAI) was significantly lower in short-stemmed cultivars (Figure 18 and Table 14) while average leaf photosynthetic rate was significantly greater (about 12%) compared to tall cassava (Figure 19 and Table 14). In these trials light interception was less in short-stemmed cassava planted at 10,000 plant ha-1 that indicated less canopy photosynthesis per unit land surface area that could be improved by increasing plant densities. Since the difference in photosynthesis was attributed mainly to nonstomatal factors, ie. biochemical and anatomical features within measophyll (El-Sharkawy and De Tafur, 2010) , selection for higher activities of key photosynthetic enzymes, ie. Rubisco and PEPC, might lead to greater yield in short-stemmed cultivars. The powerful molecular biology tools and genomics research, if properly focused toward practical applications, might further enhance progress in breeding and selection of more efficient short cassava cultivars via identification, mapping and transferring specific genes related to leaf photosynthetic capacity (including anatomy and biochemical traits), photoassimilates translocation and sink strength of storage roots, as well as sugar synthesis and starch metabolism characteristics (Bonierbale et al., 2003; Raines, 2006, 2011). A significant advance in that direction is the recent determination of the complete sequence and organization/structure of the chloroplast genome of cassava, which is the first published genome sequence of a member of the family Euphorbiaceae (Daniell et al., 2008). However, the implications of these findings for the process of cassava photosynthesis have yet to be investigated. Moreover, since significant yield losses result from several arthropod pests feeding on cassava storage roots, leaves, and/or stems, particularly members of Lepidoptera, Diptera, and Hemiptera, chloroplast genetic manipulations might help in alleviating this problem through the use of a high-dosage strategy that can kill both susceptible and resistant insects to Bacillus thuringiensis toxins. Thus, the chloroplast genome sequence may provides the necessary information for the design and construction of an efficient chloroplast vector system (Daniell et al. 2008).


189

Table 14 . Comparative productivity and leaf photosynthesis in different plant types grown under rain-fed conditions at CIAT-Quilichao Experiment Station, Cauca Department, Colombia. Data are means of two years (1994/1995, 1995/1996). Seasonal leaf photosynthesis was determined in upper canopy leaves from 3 to 8 month after planting by portable CO2 infrared gas analyser during dry periods. Final harvests were conducted 10 months after planting. Note: the higher harvest index (HI) values due to the much lower dry shoot biomass in short-stemmed cultivars. (El-Sharkawy and De Tafur, 2010) Cultivar

Tall – High LAI CG 402 -11 CM 4574 – 7 MBra 110 MMal 48 CM 507 – 37 CM 4729 – 4 SG 536 – 1 Mean Short – Low LAI MCol 22 MBra 900 CG 1141 – 1 CG 1420 – 1 CM 2766 – 5 MPan 51 CM 3294 – 4 SG 107 – 35 Mean LSD 5% between groups means

Seasonal LAI

Seasonal Pn µmol CO2 m-2 s-1

Final dry root yield tha-1

Final dry shoot biomass tha-1

Final total biomass tha-1

HI Harvest index

3.13 2.91 2.46 2.82 2.41 2.10 2.10 2.56

15.68 14.45 17.18 15.89 15.61 17.16 16.58 16.08

11.9 14.5 11.4 17.2 12.4 13.0 9.2 12.8

8.0 5.8 7.6 7.3 4.7 5.5 3.7 6.09

19.9 20.3 19.0 24.5 17.1 18.5 12.9 18.89

0.60 0.71 0.60 0.70 0.73 0.70 0.71 0.68

1.93 1.86 1.85 1.77 1.66 1.63 1.58 1.51 1.72 0.4

17.54 21.44 17.23 17.93 15.59 17.86 18.38 18.18 18.02 1.75

8.5 8.5 12.7 9.0 10.1 8.7 10.4 11.2 9.89 1.3

2.5 2.7 2.4 2.2 3.2 4.9 3.3 4.1 3.16 1.1

11.0 11.2 15.1 11.2 13.3 13.6 13.7 15.3 13.05 1.8

0.77 0.76 0.84 0.80 0.76 0.64 0.76 0.73 0.76

Nutrient use efficiency in terms of dry root production was significantly greater in shortstemmed cultivars than values in tall ones (Table 15 ).Thus, breeding for short-stemmed cultivars is warranted since resource-limited cassava producers rarely use chemical fertilizers and most cassava production occurs in marginal low-fertility soils. Table 15. Nutrient use efficiency for root production at 10 months after planting [kg(dry root) kg–1(total nutrient uptake)] for groups of tall and short cassava cultivars. Means of two years (1994/1995, 1995/1996). Source: (Adopted from El-Sharkawy et al. 1998a) Group Tall Short LSD 5 %

N 110 131 17

P 715 885 85

K 132 161 22

Ca 347 430 77

Mg 589 669 91

190

Mabrouk A. El-Sharkawy, Sara M. de Tafur and Yamel Lopez Tall - High LAI Mean 2.56

LAI

3

2

1

0 Short - Low LAI Mean 1.72

LAI

3

2

1

0 0

50

100

150

200

250

300

350

DAYS AFTER PLANTING

Figure 18. Time course of average LAI in tall- and short-stemmed cassava cultivars in the first growing cycle. Calculated LAI values were determined nondestructively by light-sensing canopy analyser based on differences between readings of light incidence above canopy and those recorded below the canopy at soil level. Values are means of all cultivars within each group. Bars are ± SD. Note: (a) the drop in mean LAI between 110- 150 DAP due to a prolonged drought; (b) the steeper decline in LAI in short cultivars from 200 to 300 DAP in contrast with trends in tall cultivars. Source: El-Sharkawy and De Tafur, 2010.

Combined to this cultivar trait, early bulking (early storage root formation and filling) should be selected for when cultivars are grown in water-limited environments (El-Sharkawy et al., 1998a, El-Sharkawy 2006b; Kamau et al., 2011). Moreover, longer leaf life (better leaf retention) and resistance to pests and diseases should be combined with short-stem trait.


191

PN [µmol (CO2 )m-2 s-1 ]

25

20

15

10 Tall - High LAI Mean 16.08 5

0

PN [µmol (CO2) m-2 s-1]

25

20

15

10

Short - Low LAI Mean 18.02

5

0 40

60

80

100

120

140

160

180

200

DAYS AFTER PLANTING

Figure 19. Time course of upper canopy leaf photosynthetic rate of tall- and short-stemmed cassava cultivars. Rates were determined in the field using portable infrared gas analysers and leaf chambers at solar PAR irradiance greater than 1000 µmol m-2 s-1. Data are means of all cultivars within each plant type group. Bars are ± SD. Note: (a) the low rates near 120 DAP coincided with a long drought period; (2) the higher rates in short-stemmed cultivars after recovery from water stress, as compared to tall cultivars. Source: El-Sharkawy and De Tafur, 2010.

SELECTION FOR PLANT TRAITS UNDERPENNING DROUGHT TOLERANCE IN CASSAVA Biochemical, Physiological and Morphological Traits Associated with Drought Tolerance Under stressful environments in the seasonally dry and semi-arid tropics, cassava possesses some inherent adaptive mechanisms for tolerance. Most important is the remarkable stomatal sensitivity to changes in atmospheric humidity as well as changes in soil water. By

192


closing stomata under water stress, the leaf remains hydrated and photosynthetically active, albeit at reduced rates, over most of its life span. Coupled with this stress avoidance mechanism is the deeper rooting capacity, where stored water in two metres depth of soil could be extracted at a slow rate leading to not only the survival of the crop during dry periods exceeding three months but also to a reasonable yield with efficient use of water and nutrients. Deeper rooting capacity is another important trait for selection and breeding of improved germplasm targeted to drier zones. In addition, the leaf canopy is much reduced under prolonged stress, contributing to lower crop water consumption. When recovered from stress, cassava rapidly forms new leaves with higher photosynthetic capacity, which compensates for yield reductions due to previous prolonged stresses. Productivity over a wide range of germplasm and in different environments correlates with upper canopy leaf photosynthesis as measured in the field, and the relationship is due mainly to non-stomatal factors, ie, biochemical/anatomical leaf traits. Among these leaf traits are elevated activities of the C4 enzyme PEPC. Leaf area duration under stress, coupled with host plant resistances/tolerances to pests and diseases, is a critical trait, since yield correlates with plant leaf retention capacity (Lenis et al, 2006). Drought tolerance in cassava can be assessed by measurement of the associated physiological traits. The traits underlying such high potential productivity in excess of 15t ha−1 oven-dried roots with 85 percent starch in 10–12 months under prolonged water stress include: (i) leaf conductance; (ii) deeper rooting capacity or greater extent of the fibrous root network; (iii) high leaf photosynthetic potential comparable with those in efficient C4 crops (rates under high humidity, wet soil, high leaf temperature and high solar radiation exceed 40μmol CO2 m−2 s−1); (iv) long leaf life (> 60 days), the leaves remaining active during most of their life span; (v) a sustainable leaf canopy that optimises light interception during a significant portion of the growth cycle; (vi) a high HI (> 0.5) coupled with a strong root sink (larger number of storage roots per plant); and (vii) high weights of fresh root and fresh foliage.

Germplasm Evaluationin in the Field for Selecting Drought-Tolerant Cultivars Results also point to the importance of field research versus studies carried out on plants grown indoors in greenhouses and growth cabinets (El-Sharkawy 2004, 2005, 2006a,b, 2007, 2010). Well-designed and planned field experiments to determine relevant physiological traits are the most cost-effective way of evaluating drought tolerance in cassava. Appropriate sites are seasonally dry and semi-arid regions with 600-800 mm of rainfall distributed over 5-8 months of the year such as in Northern Colombia, Pacific Coasts of Ecuador and Peru, Northeastern Brazil. Cassava requires three months of rainfall for establishment of the crop, after which, the crop tolerates extended water deficits greater than 3 months. Sites in Colombia used in conducting the research summarized above for evaluation and selection of drought tolerant cultivars under natural precipitation (De Tafur et al., 1997a,b; El-Sharkawy 1993, 2007, 2010; El-Sharkawy et al., 1990; El-Sharkawy and Cock, 1990) included:

Eco-Physiological Research for Breeding Improved Cassava Cultivars …   

193

Semi-arid site at Rio Hacha, Guajira Department: latitude: 11°32' N ; longitude: 72°54' W. Seasonally dry site at Santo Tomas, Atlantic Department: latitude: 10° 57' N; longitude: 74° 47' W. Seasonally dry site at Patia Valley, Cauca Department: latitude 2° 09' N; longitude: 77° 04' W.

It is noteworthy to mention that all three testing sites were small Colombian farmers fields who offered their collaboration and hospitality to the CIAT cassava research team during the course of this long research. To them we dedicate the hard-obtained achievements outlined in this chapter.

CONCLUSION The research reviewed here on cassava productivity, physiology and/or ecophysiology, and responses to environmental stresses was conducted in collaboration with a multidisciplinary team at CIAT. The Center also holds a diverse germplasm bank of the crop, which has been assembled over 40 years. The research objectives revolved around the strategy adopted for developing new technologies to enhance crop productivity in most of the edaphoclimatic zones under which cassava is cultivated, particularly stressful environments. Under favorable environments in lowland and mid-altitude tropical zones with nearoptimal climatic and edaphic conditions for the crop to realize its inherent potential, cassava is highly productive in terms of root yield and total biological biomass. For example, under trial conditions, improved germplasm can, after 10–12 months’ growth, attain >15 t/ha of oven-dried roots, containing 85% starch. The physiological mechanisms underlying such high potential productivity are: 1. High leaf photosynthetic potential, comparable with those in efficient C4 crops (assuming the following conditions are common: high humidity, adequate soil moisture, high leaf temperature, high solar radiation, and a Pn rate that exceeds 40 µmol CO2 m-2 s-1 ); 2. Long leaf life (>60 days), with the leaves remaining active for most of their life spans; 3. Sustainable leaf canopy that optimizes light interception during a significant portion of the growth cycle; and 4. High harvest index (>0.5), coupled with strong root sink (i.e., larger number of storage roots/plant). Under stressful environments in seasonally dry and semi-arid tropics, productivity is reduced, with more reduction in aboveground parts (shoots) than in storage roots (i.e., higher HI). Under these conditions, the crop possesses some inherent adaptive mechanisms for tolerance. The most important one is the remarkable stomatal sensitivity to changes in atmospheric humidity, as well as in soil water. By closing stomata under water stress, the leaf remains hydrated and photosynthetically active, although at reduced rates, over most of its life span.

194


Together with this “stress avoidance mechanism” is a deep rooting capacity that enables the plant to slowly extract water from as deep as 2 m. The crop therefore not only survives dry periods of up to 3 months long, but also produces a reasonable yield through its efficient use of water and nutrients. Moreover, leaf canopy is much reduced under prolonged stress, contributing to lower crop water consumption. When it recovers from stress, cassava rapidly forms new leaves with higher photosynthetic capacity, which compensates for yield reductions from the previous prolonged stress. Productivity over a wide range of germplasm and in different environments correlates with upper canopy leaf photosynthesis, as measured in the field. The relationship stems mostly from non-stomatal factors, that is, from biochemical and anatomical leaf traits. Among these leaf traits is the elevated activity of the C4 PEPC enzyme. Breeding programs could exploit the genetic variations found within cassava germplasm and wild Manihot for both leaf photosynthesis and enzyme activity. Leaf area duration under stress, together with host-plant resistance or tolerance of pests and diseases, is a critical trait because yield correlates with leaf retention capacity (Lenis et al., 2006). Deep rooting capacity is another important trait for selecting and breeding improved germplasm for drier zones. In cooler zones such as higher altitudes in the tropics and lowland subtropics, cassava growth is slower and the crop stays in the ground for longer time to achieve adequate yields. Under these conditions, leaf formation is slower, leaf photosynthesis is much reduced, but leaf life is longer (Irikura et al. 1979; El-Sharkawy et al., 1992a, 1993). Wide genetic variations exist for photosynthesis that may be valuable for selecting and breeding for genotypes that better tolerate cool climates. Combining enhanced leaf photosynthesis with the normally longer leaf life in cool climates may improve productivity. Selecting for medium-statured and short cassava instead of tall cassava is advantageous for saving on nutrient uptake and ensuring higher nutrient-use efficiency for root production without sacrificing potential yield. Short cultivars showed a tendency for early bulking, a favorable selectable trait for seasonally dry and semi-arid environments (El-Sharkawy et al., 1998a; El-Sharkawy, 2006b; El-Sharkawy and De Tafur, 2010) . Germplasm from the core collection was screened for tolerance of soils low in P and K, resulting in the identification of several accessions with good levels of tolerance. Results also point to the importance of field research versus greenhouse or growthchamber studies that do not calibrate for representative environments to account for acclimation factors (El-Sharkawy, 2005). Calibration becomes even more critical when data from indoor-grown plants are used to extrapolate to the field or to develop crop models.

FUTURE PROSPECTS FOR CASSAVA IMPROVEMENT Much remains to be done to further improve productivity while conserving dwindling natural resources such as water and land. Developing countries, in particular, need more support to continue with maintenance research, which aims to upgrade previous findings and technologies; contribute to sustainable agricultural, economic, and social developments; and enhance food supply to meet increasing demands under observed trends in global climate changes. Cassava is unique among staple crops in meeting these demands because of its inherent tolerance to adverse eco-edaphic conditions such as higher temperature, prolonged drought and poor soils.


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Basic research can be cost-effective, with high returns, even if slower. It can be especially successful when conducted in collaboration with multidisciplinary and/or multiinstitutional teams that follow well-planned strategies and are focused towards fulfilling a set of high priority goals and objectives. Plant physiology, biochemistry and molecular genetics are essential components in such endeavors. Since cassava is genetically heterozygous and vegetative propagated for commercial production, genetic manipulation aided by advanced biotechnology tools should further enhance progress in cassava improvement in the near future, particularly under stressful environments, as the plant is amenable to genetic transformation (Li et al., 1996; Puonti-Kaerlas, 1998). This genetic molecular approach is of paramount importance particularly for characterizing germplasm pools, selecting plants tolerant/resistant of devastating pest diseases, for food and feed biofortifications of storage roots with essential nutrients (eg, higher content of protein, carotenes, trace minerals such as iron and zinc), all are characteristics that might enhance productivity and qualities of cassava end-products (Schuch et al., 1996; Bonierbale et al., 2003; Ferriera et al., 2008; Stupak ,2008;Montagnac et al., 2009b; Nassar and Ortiz, 2010; Abhary et al., 2011; Sayre et al., 2011; Vieira et al., 2011). Cassava germplasm is widely diverse in storage root carotenes content (Iglesias et al., 1997; Ferriera et al., 2008) that are essential nutrients for provision of Vitamin A in human diets. Breeding programs of national and international research institutions are currently embarked on developing improved cultivars rich in these compounds. It is anticipated that when released these improved cultivars rich in pro-Vitamin A carotenes, will be also drought tolerant/resistant for its potential cultivation in droughtstricken tropical/subtropical regions where cassava intake is quite high. Worldwide research effort for biofortification of staple food crops via breeding improved cultivars was launched in the past two decades (Welch and Graham, 2002). International research and development organizations, donor agencies, and private sectors should take leading roles in financing and supporting basic research efforts, particularly those oriented towards serving the technological needs of resource-poor developing countries. Without the steady funding by only a few international donors during the 1940s and 1950s, the so-called ´Green Revolution´ of the 1960s, which had saved many developing countries from famine, would probably never have happened (Wortman,1981). The Green Revolution´s research was mainly conceived and conducted by few committed international researchers working in close collaboration with national programs across continents (Borlaug, 1983). The current CGIAR policy of conducting short-term research projects which in most cases are serving the self-interest of those involved, instead of mission-and-team-oriented and problemsolving core research, is counterproductive and should be reversed to avoid waste of resources and time (eg, Okogbenin, 2010). Scientists as well as research managers must be hold accountable for delivering the goods expected by the tax-providing public, donors, and developmental agencies.

ACKNOWLEDGMENTS The authors are indebted to the many colleagues, associates, students, workers and secretaries at CIAT whose dedication and support were crucial in realising the research achievements highlighted in this review article. The many useful reprints received from G Bowes, V Gutschick, P Jarvis, C Körner, R Lösch, F Meinzer, O Ueno and P Westhoff were

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appreciated. We thank MB Kirkham for the copy of her invaluable text book on Principles of soil and plant water relations. Useful comments on an original version were received from Drs D Laing (former Deputy Director General for Research at CIAT) and JD Hesketh (USDA/ARS, University of Illinois, Urbana, Illinois, USA). We are grateful to Farah ElSharkawy Navarro for her invaluable help in searching the internet for pertinent information, assembling the numerous data in tables and figures scattered in various published and unpublished documents, and in typing the manuscript. The logistic and financial supports from various governments and research-funding organisations including Australia, Brazil, Colombia, Germany, Switzerland and the USA, were crucial in conducting this research, which is targeted to the needs of some of the poorest farmers in developing countries.

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