Transgenic banana for Africa

Leena Tripathi, l.tripathi@cgiar.org

Banana (Musa spp.) are one of the most important food crops after maize, rice, wheat, and cassava. Annual production in the world is estimated at 130 million t, nearly one-third of it grown in sub-Saharan Africa, where the crop provides more than 25% of the food energy requirements for over 100 million people. East Africa is the region that produces and consumes the most banana in Africa. Uganda is the world’s second largest producer after India, with a total of about 10 million t.

Banana plantation damaged by Xanthomonas wilt. Photo by IITA.
Banana plantation damaged by Xanthomonas wilt. Photo by IITA.

The banana Xanthomonas wilt (BXW) disease caused by the bacterium Xanthomonas campestris pv. musacearum (Xcm) was first reported about 40 years ago in Ethiopia on Ensete spp., a close relative of banana. Outside Ethiopia, BXW was first identified in Uganda in 2001, subsequently in the DR Congo, Rwanda, Kenya, Tanzania, and Burundi. The disease is highly contagious and is spread plant-to-plant through the use of contaminated agricultural implements. It is also carried by insects that feed on male buds, and is present on plant material, including infected debris. The rapid spread of the disease has endangered the livelihoods of millions of farmers who rely on banana for staple food and cash.

Infection by Xcm results in the yellowing and wilting of leaves, uneven and premature ripening of fruits, and yellowish and dark brown scars in the pulp. Infected plants eventually wither and die. The pathogen infects all varieties, including East African Highland Banana (EAHB) and exotic types, resulting in annual losses of over US$500 million across East and Central Africa.

Options for BXW control using chemicals, biocontrol agents, or resistant cultivars are not available. Although BXW can be managed by following phytosaniary practices, including cutting and burying infected plants, restricting the movement of banana materials from BXW-affected areas, decapitating male buds, and using “clean” tools, the adoption of such practices has been inconsistent. They are labor-intensive and farmers believe that debudding affects the fruit quality.

The use of disease-resistant cultivars has been an effective and economically viable strategy for managing plant diseases. However, resistance to BXW has not been found in any banana cultivar. Even if resistant germplasm is identified, conventional banana breeding to transfer resistance to farmer-preferred cultivars is a difficult and lengthy process because of the sterility of most cultivars and also the long generation times.

Transgenic technologies that facilitate the transfer of useful genes across species have been shown to offer numerous advantages to avoid the natural delays and problems in breeding banana. They provide a cost-effective method to develop varieties resistant to BXW. Transgenic plants expressing the Hypersensitive Response Assisting Protein (Hrap) or Plant Ferredoxin Like Protein (Pflp) gene originating from sweet pepper (Capsicum annuum) has been shown to offer effective resistance to related Xanthomonas strains.

Plants established in confined field trial 5 months after planting. Source: L. Tripathi, IITA.
Plants established in confined field trial 5 months after planting. Source: L. Tripathi, IITA.

IITA, in partnership with the National Agricultural Research Organization (NARO)-Uganda and the African Agriculture Technology Foundation (AATF), has developed transgenic banana expressing the Hrap or Pflp gene using embryogenic cell suspensions or meristematic tissues of four banana cultivars, Sukali Ndiizi, Mpologoma, Nakinyika, and Pisang Awak. More than 300 putatively transformed plants were regenerated and validated via PCR assay and Southern blot. Of these, 65 transgenic plants have exhibited strong resistance to BXW in the laboratory and screenhouse tests. The plants did not exhibit any differences from their nontransformed controls, suggesting that the constitutive expression of these genes has no effect on plant physiology or other agronomic traits.

The 65 resistant lines were planted in a confined field trial in October 2010 at the National Agriculture Research Laboratories (NARL), Kawanda, Uganda, after approval was obtained from the National Biosafety Committee. These transgenic lines are under evaluation for disease resistance and agronomic performance in field conditions. The transgenic lines are slated for environmental and food safety assessment in compliance with Uganda’s biosafety regulations, and procedures for risk assessment and management, and seed registration and release. After completing the necessary biosafety validation and receiving approval from the Biosafety Committee, the Xcm-resistant cultivars are expected to be deregulated for cultivation in farmers’ fields in Uganda.

We plan to stack the Pflp and Hrap genes in the same cultivars to enhance the durability of resistance against Xcm. We have developed more than 500 transgenic lines with the double genes construct (pBI-HRAP-PFLP) which are being evaluated for disease resistance under contained screenhouse conditions.

This technology may also provide effective control of other bacterial diseases such as moko or blood disease, of banana occurring in other parts of the world. The elicitor-induced resistance could be a very useful strategy for developing broad-spectrum resistance. The elicitor is a protein secreted by pathogens that induce resistance. The transgenic banana carrying these genes may also display resistance to fungal diseases such as black sigatoka and Fusarium wilt. Experiments on this are being conducted in our lab in Uganda.

Confined field trial of banana plants. Source: L. Tripathi, IITA.
Confined field trial of banana plants. Source: L. Tripathi, IITA.

We are also planning to stack genes for resistance to Xcm and nematodes into one line to produce cultivars with dual resistance that would tackle two of the most important production constraints in Eastern Africa.

The development of Xcm-resistant banana using the transgenic approach is a significant technological advance that will increase the available arsenal of weapons to fight the BXW epidemic and save livelihoods in Africa. It can become a high-value product for farmers.

This research is supported by the Gatsby Charitable Foundation, AATF, and USAID.

Note: The Pflp and Hrap genes are owned by Taiwan’s Academia Sinica, the patent holder. IITA has negotiated a royalty-free license through the AATF for access to these genes for use in the commercial production of BXW-resistant banana varieties in sub-Saharan Africa.

New approaches to assessing soil conservation options

Birte Junge, Birte.Junge@web.de

Land degradation has become a global concern as it affects the environment, agronomic productivity, food security, and quality of life (Eswaran et al. 2001). Due to rapid population growth, land use has been intensified to cover the increased demand for food. This often results in the loss of soil and nutrients when land is not used properly.

Change in village area, Badume, Kano State, Nigeria. Image from B. Junge.
Change in village area, Badume, Kano State, Nigeria. Image from B. Junge.

Processes that degrade soil include the loss of topsoil by the action of water or wind; chemical deterioration, such as nutrient depletion; physical degradation, such as compaction; and biological deterioration of the natural resource which includes, among others, the reduction of soil biodiversity (Lal 2001).

In Nigeria, where the population dramatically increased from 115 million in 1991 to 140 million in 2006 (FRN 2007), human-induced soil degradation is a common phenomenon: its severity is low for 37.5% of the area (342,917 km2), moderate for 4.3% (39,440 km2), high for 26.3% (240,495 km2), and very high for 27.9% (255,167 km2) (FAO 2005).

Soil erosion is the most widespread type of soil degradation in the country and has long been recognized as a serious problem (Stamp 1938). In 1989, 693,000 km2 were already characterized by runoff-induced soil loss in the south. In the north, 231,000 km2 were degraded, mainly by wind erosion. Sheet erosion is observed all over the country. Rill and gully erosion are common in the east and along rivers in northern Nigeria (Ologe 1978, Igbozurike et al. 1989).

Soil erosion degrades the natural resource base, resulting in the loss of land for farming or grazing animals, as well as off-site problems, such as the sedimentation of dams. Reduced agricultural production, food insecurity, low income for the rural population, and poverty are some of the consequences. Thus, more emphasis has to be put on avoiding soil loss through improved management to conserve the natural resources for today and the future.

Monitoring land use intensification and soil degradation
A German Federal Ministry for Economic Cooperation and Development/German Society for Technical Cooperation (BMZ/GTZ) project from 2005 to 2008 included the study of soil erosion and its causes using different methodologies, such as remote sensing and the geographic information system (GIS) technology. Aerial photos and satellite images produced at different dates and scales and GIS can be used to obtain key information on environmental resources and their degradation (Oluseyi 2002).

Remote sensing data: aerial photograph, 1962/1981. Image from B. Junge.
Remote sensing data: aerial photograph, 1962/1981. Image from B. Junge.
Remote sensing data: IKONOS, 2005. Image from B. Junge.
Remote sensing data: IKONOS, 2005. Image from B. Junge.
Remote sensing data: Quickbird, 2005/06/07. Image from B. Junge.
Remote sensing data: Quickbird, 2005/06/07. Image from B. Junge.

Badume (12°19’N 8°31’E), Kayawa (11°22’N 7°20’E), Gadza (8°98’N 6°00’E), and Eglime (7°08’N 1°67’E), villages located in different agroecological zones of Nigeria and Bénin, were selected as study sites. Historical aerial photos (from 1962, 1981, or 1982) and recent satellite images (IKONOS imagery with spatial resolution of 1 m from 2000; and QuickBird imagery with spatial resolution of 0.6 m from 2005 to 2007) were analyzed to study the change in land use/land cover (LULC) and soil degradation in these places.

A manual interpretation approach was used to identify the LULC classes of the study areas. Verification was done in the field with local farmers and by using the global positioning system (GPS) to obtain the accurate location of the former and present village boundaries and to get point data for the LULC classes. In addition, the average growth rate of several gullies in each village was measured for use in forecasting possible expansion in the future.

Change in LULC
The interpretation of historical and recent remote sensing data showed that the area of all settlements expanded within the last decades and years. For example, the village of Badume increased from 0.9 ha (1962) to 4.3 ha (2006), an expansion of 3.4 ha in 44 years. The rate of increase was slower in former times than in recent years. This reflects the recent rise in population.
The area in use around the pilot villages also expanded within the period considered. For example, the area of Kayawa increased from 166.8 ha to about 438.6 ha, an expansion of 272.6 ha in 44 years. The rate of increase was much higher in former times than in recent years, with more of the uncultivated land available around the settlements being converted into farmland. No uncultivated land was/is available for expansion any longer, since the borders with neighboring villages had been reached.

The areas covered with trees or shrubs generally decreased in the pilot villages. For instance, in Gadza, the area decreased by 4.8 ha from 2000 to 2005. Causes are conversion into farmland, but also the uncontrolled exploitation of these areas for fuelwood (Odihi 2003), timber (Okoro 1990), or logging (Omo-Irabo and Odunyemi 2007). The reduction or elimination of fallow was especially observable in Badume and Kayawa, but not so severe in Gadza where fallow still covered a certain percentage of the village area. The reason might be the production of cash crops in inland valleys—the “fadama”—which ensures a certain income. Uncultivated areas in the surroundings of the fields also decreased during the period under consideration. Having less land is a constraint to livestock production for the Fulani (migrant farmers) who need to graze their animals. Conflicts between arable farmers and pastoralists already take place in the pilot villages because of competition for limited land resources.

Change in soil degradation
The study sites located in the Sudan and northern Guinea savanna were characterized by sheet erosion. In Badume, the eroded area near the river sites increased from 11.6 ha in 2000 to 12.3 ha in 2006. In Kayawa, sheet erosion was a big problem, especially in the western part of the village area. Land characterized by sheet erosion (7.9 ha) was already visible on the photo in 1962 and increased to 32.3 ha in 2006.

Gully erosion, Badume, Kano State, Nigeria, 2000 (left) and 2006 (right).
Gully erosion, Badume, Kano State, Nigeria, 2000 (left) and 2006 (right).

Erosion gullies were also detected along the rivers in the uncultivated area of Badume and Kayawa. The study of two gullies located northeast of the village area of Badume revealed that the degraded land increased from 37.9 ha (2000) to 45.1 ha (2006). In Kayawa, gully erosion also increased. In 1962, about 1.5 ha of the village area was destroyed by gullies. This area increased to 13.1 ha in 2000 and to 15.5 ha in 2006. Reasons include the high impact of the rain at the beginning of the wet season after the total removal of crop residues during the dry season (Odunze 2003) and the formation of ridges along the slope that increase the velocity of runoff and hence, the removal of topsoil (Lal 1995). The herds of cattle that graze on the land or pass it on their way to watering pits or other pastures are very destructive. The frequent shortage of grass, overgrazing, and trampling by animals result in a sparse vegetation cover that exposes bare soil and accelerates the formation of runoff and the removal of topsoil (Azeza and Omeji 1985).

The uncultivated area surrounding Gadza was characterized by a huge network of deeply worn animal tracks mainly located between a Fulani settlement and the Gadza River. These pathways were carved to different extents, depending on their location and the frequency of animal passage. During heavy rain, the transport of runoff and sediment was observed, resulting in the removal of fine particles and the accumulation of sand and gravel. In 2000, about 17 km of track-gullies already existed; this increased to 83.7 km in 2005. This immense increase of 66.7 km in 5 years might be caused by the higher resolution of the recent QuickBird images which facilitated the detection of small tracks. Another reason might be the increase in the number of cattle causing these linear erosion features.

No erosion gullies were detected on the aerial photograph of Eglime made in 1982. This might be caused by the small scale of the picture and its bad quality, as elderly farmers remember the presence of some erosion features from their childhood. In 2000, gullies with a total length of 4.4 km were observed and this total increased to 42.2 km in 2007. Reasons for this big increase are the good quality of recent images and the change in LULC. The farm area has increased within the last decades with the promotion of cotton cultivation. This crop is known for its low canopy cover and, hence, for reduced protection of the soil surface against the impact of rain drops (Junge 2004).

Based on calculated gully growth rates, the estimation of possible future expansion of areas degraded by gully erosion showed an increase for all pilot villages, but to different extents. In Badume, 3.0 ha and in Kayawa, 4.7 ha are expected to be eroded in 10 years. The gullies in Gadza, developed from animal tracks, will cover an area of about 1.2 ha in the following decade, and in Eglime, 2.3 ha of land will be lost to gully erosion.

Using remote sensing and GIS
The interpretation of remote sensing data produced at different dates and of pilot villages in different agroecological zones of Nigeria and Bénin has shown that the village area, farmland, and settlement expanded at all sites. Farmland increased at the expense of areas usually covered with forest and shrubs, fallow, and grazing land.

The consequences of this land use intensification were detected in all study sites in the form of sheet and gully erosion. The removal of soil including organic matter and nutrients inevitably resulted in the decrease of arable and grazing land, reduced production of crops, meat, and milk products, and reduced income for the local farmers. If no measures to conserve the natural resources are implemented in the near future, the increase in soil deterioration will continue. Land scarcity will follow with related conflicts between the users, food insecurity, and poverty.

The generation of an environmental database and the mapping of LULC and existing soil degradation are the bases for improved natural resource management. Detailed surveys are therefore recommended to intensify the land use inventory to generate an environmental database in Nigeria. This would include information on present LULC and the extent of soil degradation.

Land use plans should also be developed illustrating the areas used for agriculture (farming and pasturing), urban development, industry, and commercial purpose. Information of these kinds are a useful tool for policymakers and land use planners and would contribute to an enhanced management of the environment in Nigeria and Bénin.

References
Azeza, M.I. and M.U. Omeje. 1985. Soil erosion control measures in the Sahel. In Proceedings of Conference National Workshop on Ecological Disasters in Nigeria: Drought and Desertification. 9–12 December 1985. Kano, Nigeria. pp. 377–379.

Eswaran, H., R. Lal, and P.F. Reich. 2001. Land degradation: an overview. In Responses to Land Degradation, edited by E.M. Bridges, I.D. Hannam, L.R. Oldeman, F.W.T. Pening de Vries, S.J. Scherr, and S. Sompatpanit. Proceedings of the 2nd International Conference on Land Degradation and Desertification, Khon Kaen, January 1999. New Delhi: Oxford Press. http://soils.usda.gov/use/
FAO, AGL. 2005. Global Assessment of the Status of Human-Induced Soil Degradation (GLASOD). http://www.fao.org/landandwater/agll/glasod/glasodmaps.jsp.

FRN (Federal Republic of Nigeria). 2007. Official Gazette: Legal Notice on publication of the details of the breakdown of the National and State provisional totals 2006 Census. Government Notice Nr. 21. Nr. 24. Vol. 94. Lagos.

Igbozurike, U.M., D.U.U. Okali, and A.T. Salau. 1989. Profile on Nigeria: Land Degradation. Report submitted to Commonwealth Secretariat, London. Friedrich Ebert Foundation, Lagos, and Nigerian Environmental Study/Action Team (NEST), Ibadan, 48 pp.

Junge, B. 2004. Die Boeden des oberen Oueme-Einzugsgebietes in Benin/Westafrika – Pedologie, Klassifizierung, Nutzung und Degradierung. PhD dissertation, University of Bonn, Germany. 291 pp.

Junge B., T. Alabi, K. Sonder, M.Subash, R. Abaidoo, D. Chikoye, and K. Stahr. 2009. Use of remote sensing and GIS for monitoring changes of land use/land cover and environmental degradation in different agroecological zones of West Africa. Int. J. Remote Sensing. (in press)
Lal, R. 1995. Sustainable management of soil resources in the humid tropics. United Nations University Press. New York, USA. 593 pp.

Lal, R. 2001. Soil Degradation by Erosion. Land Degradation and Development 12:519–539.
Odihi, J. 2003. Deforestation in afforestation priority zone in Sudano-Sahelian Nigeria. Applied Geography 23(4):227–259.

Odunze A.C. 2003. Northern Guinea savanna of Nigeria and rainfall properties for erosion control. African Soils 33:73–116.

Okoro, S.P.A. 1990. Status of forest resources of Nigeria. Paper presented at the 20th Annual Conference of the Forestry Association of Nigeria. 25–30 November 1990, Katsina, Nigeria. Forestry Association of Nigeria. 20 pp.

Ologe, K.O. 1978. A quick preliminary survey of soil erosion in northwestern Nigeria. Report for the Land Resource Division of the Federal Department of Agriculture, Kaduna. 18 pp.

Oluseyi, F.O. 2002. Integration of remote sensing data and field models of in-situ data in a GIS for environmental sensivity index mapping; a Nigerian example. Available online at: http://www.isprs.org/commission4/proceedings02/pdfpapers/464.pdf (accessed 28 July 2008).

Omo-Irabo, O.O. and K. Odunyemi. 2007. A hybrid image classification approach for the systematic analysis of land cover (LC) changes in the Niger Delta Region. Available online at: http://www.itc.nl/ISSDQ2007/proceedings/Session%202%20Spatial%20Statistics/paper%20omoleomo.pdf (accessed 27 July 2008).

Stamp, L.D. 1938. Land utilization and soil erosion in Nigeria. Geographical Review 28:32–45.
Tappan, G. and M. Cushing. 2004. Use of SCL-Off Landat image data for monitoring land use/ land cover trends in West Africa. USGS EROS Data Center, Sioux Falls, SD, USA. 11 pp.

DNA barcodes for pathogens of African food crops

Lava Kumar, L.kumar@cgiar.org and Kamal Sharma, k.sharma@cgiar.org

Diagnostic tools play an important role in the accurate and timely identification of the pathogens involved in disease etiology, also in disease surveillance, the development of host plant resistance, quarantine monitoring, and support safe conservation and the exchange of germplasm. Detailed knowledge of pathogen population structure and genetic diversity is a prerequisite to developing unambiguous diagnostic tools and is critical in establishing disease management tactics.

Severe anthracnose symptoms on cassava stem. Photo by R. Bandyophadyay, IITA.
Severe anthracnose symptoms on cassava stem. Photo by R. Bandyophadyay, IITA.

Increasingly, modern diagnostic tools are being based on the DNA characteristics of the pathogen as they are neutral to growth stage and environment; offer adequate diversity to distinguish species, strains, substrains, isolates, and even individuals; and offer convenience of detection using modern bio-techniques such as polymerase chain reaction (PCR).

At IITA, we undertook a new initiative to characterize pathogen populations and recognize unique stretches of sequences—known as ”DNA barcodes”—that can be used as genetic markers for the rapid diagnosis of the pathogens and pests affecting the African food crops on which we work. DNA barcodes, otherwise also known as DNA markers or DNA fingerprints, are essentially a short stretch of nucleotide sequences that aid in the specific identification of species strains or substrains. They are used to resolve pathogen taxonomy and phylogeny.

The work focuses on economically important fungal, viral, and bacterial pathogens, insects, and nematodes. The information is used to gain ”barcode” designation in global sequence databases such as BOLD (the barcode of life data system) or NCBI (National Center for Biotechnology Initiative), and to assemble these into a database for public access.

This approach—a combination of conventional biology, biotechnology, and bioinformatics—involves the selection of targets, amplification of target genes using universal or generic primers, sequencing of target genes and identification of unique barcodes, and development of PCR-based diagnostics for specific detection of barcodes. This approach is particularly useful in identifying pathogens that are difficult to distinguish either by morphology or other properties. It offers high accuracy in identifying quarantine pathogens and reduces the risk of spread. In addition to diagnosis, it also contributes to the fundamental understanding of pathogen phylogeography and relationship with host and contributes to the development of management tactics.

Clustering of 25 yam isolates based on rDNA sequences. Courtesy of Lava Kumar, IITA.
Clustering of 25 yam isolates based on rDNA sequences. Courtesy of Lava Kumar, IITA.

We are using this approach to characterize the fungal pathogen(s) causing anthracnose—the most destructive disease of yam and cassava in West Africa. The disease causes severe yield losses in both crops and often kills the plant. The causal fungus, Colletotrichum gloeosporioides Penz., is widespread in West Africa. We identified various isolates of this fungus differing in morphology, growth characters, and pathogenicity, then investigated their genetic relatedness and diversity through molecular analysis of a set of 25 reference isolates (17 from yam and 8 from cassava) using multilocus gene targets. They were grouped into spot (S) and blight (B) isolates based on symptoms they induce. Both types of isolates infect yam, but only B isolates infect cassava. We assessed the genetic diversity in these isolates by nucleotide sequencing and cluster analysis of the ~540 base pair (bp) nuclear ribosomal internal transcribed spacer region (ITS1, ITS2 and the 5.8S gene) and partial gene sequences of actin (~240 bp) and histone (~370 bp).

Phylogenetic cluster analysis grouped the 25 isolates into two major clades (a clade is a group that shares features from a common ancestor) and two subclades within the major clades. Both the S and B isolates were distributed between the two clades (see figure). All the isolates in clade 1 were unique to yam. Seven of these isolates (YA08-1, YA08-2, YA08-3, YA08-4, YA08-7, Y-83, Y-84) formed a genetically distinct lineage, indicating that they could be new strains unique to yam. Isolates in clade 2 infect both cassava and yam, suggesting their capability to infect a wide range of plants. It is plausible that clade 2 isolates could be those most frequently occurring on yam and cassava because of their ability to survive on weeds and other crops. We recognized unique sequence motifs and designed diagnostic PCR primers directly from infected plant tissues for the specific amplification of C. gloeosporioides infecting yam and cassava.

Gray leaf spot lesions in maize. Photo by A. Aregbesola, IITA.
Gray leaf spot lesions in maize. Photo by A. Aregbesola, IITA.

Using a similar approach, we characterized the fungal agent associated with gray leaf spot (GLS), the most destructive disease of maize. We found that GLS in Nigeria is caused by a distinct species of Cercospora, but not C. zeae-maydis, a previous conclusion derived from conventional analysis. This work, in addition to confirming the GLS etiology, allowed us to establish a unique set of primers for the specific identification of the GLS pathogen prevalent in Nigeria.

Through comparative genomics, we identified common genome regions in cassava mosaic begomoviruses occurring in sub-Saharan Africa. We developed a simple multiplex PCR assay that can detect all the major viruses in cassava mosaic disease etiology. This test has been adopted for virus indexing of cassava propagated in vitro.

To aid us in diagnostics research, we developed a simple and cost-effective procedure suitable for extraction of DNA from seeds, leaves, stems, tubers, and even roots. The resultant DNA is suitable for PCR-based diagnoses of fungi, bacteria, and viruses in the infected tissues of a wide range of plant species. It is handy for the quarantine monitoring of germplasm. We are establishing a repository of diagnostic protocols in an approach we call the ”Diagnostic Basket®” and will make it available to users.

Barcodes and diagnostic tools provide a solid base for the understanding of the taxonomy and diversity of pathogens infecting African food crops.

African yam bean: a food security crop?

Daniel Adewale, d.adewale@cgiar.org

Read the Ukranian translation by Martha Ruszkowski

Diversity in color, color pattern, structure, texture, brilliance, etc. of African yam bean seeds. Photo by D. Adewale, IITA.
Diversity in color, color pattern, structure, texture, brilliance, etc. of African yam bean seeds. Photo by D. Adewale, IITA.

Biodiversity assures the evolutionary continuity of species. The collection and conservation of diversity within species are a safeguard against the loss of germplasm. They provide a buffer against environmental threats and assure continual and sustainable productivity. Global food security is becoming shaky with increasing dependence on a few major staple crops. This has resulted in an alarming reduction not only in crop diversity but also in the variability within crops.

The conservation and maintenance of agrobiodiversity of neglected and underutilized plant species such as African yam bean (AYB) in seed banks aim at contributing to food security and preventing a potential food crisis. Increasing the use of underutilized crops is one of the better ways to reduce nutritional, environmental, and financial vulnerability in times of change (Jaenicke and Pasiecznik 2009); their contribution to food security is unquestionably significant (Naylor et al. 2004, Oniang’o et al. 2006). Among other things, the consumption of a broader range of plant species ensures good health and nutrition, income generation, and ecological sustainability.

Potentials of African yam bean
The plant (Sphenostylis stenocarpa) is one of the most important tuberous legumes of tropical Africa. It is usually cultivated as a secondary crop with yam in Ghana and Nigeria. A few farmers who still hold some seed stocks, especially the white with black-eye pattern, plant it at the base of yam mounds in June or July. The crop flourishes and takes over the stakes from senescing yam. It flowers and begins to set fruits from late September and October. The large bright purple flowers result in long linear pods that could house about 20 seeds.

The seed grains and the tubers are the two major organs of immense economic importance as food for Africans. This indigenous crop has huge potential for food security in Africa. However, there are cultural and regional preferences. In West Africa, the seeds are preferred to the tubers but the tubers are relished in East and Central Africa (Potter 1992). The crop replaces cowpea in some parts of southwestern Nigeria (Okpara and Omaliko (1995). Researchers (Uguru and Madukaife 2001) who did a nutritional evaluation of 44 genotypes of AYB reported that the crop is well balanced in essential amino acids and has a higher amino acid content than pigeon pea, cowpea, and bambara groundnut.

Tuber yield per stand of AYB accession TSs96 at Ibadan, 2006. Photo by D. Adewale, IITA.
Tuber yield per stand of AYB accession TSs96 at Ibadan, 2006. Photo by D. Adewale, IITA.

Apart from the use of soybean as an alternative to animal protein, protein from other plant sources is not often exploited. The protein content in AYB grains ranged between 21 and 29% and in the tubers it is about 2 to 3 times the amount in potatoes (Uguru and Madukaife 2001, Okigbo 1973). AYB produces an appreciable yield under diverse environmental conditions (Anochili 1984, Schippers 2000). Another positive contribution of the crop to food security is the identification of the presence of lectin in the seeds, which could be a potent biological control for most leguminous pests.

Biodiversity
Although the vast genetic and economic potentials of AYB have been recognized, especially in reducing malnutrition among Africans, the crop has not received adequate research attention. Up to now, it is classified as a neglected underutilized species or NUS (Bioversity 2009). Devos et al. (1980) stressed that the danger of losing essential germplasm hangs over all cultivated food crop species in tropical Africa, especially those not receiving research attention. The quantity and availability of AYB germplasm is decreasing with time. At one time, Klu et al. (2001) had speculated that the crop was nearing extinction; its inherent ability to adapt to diverse environments (Anochili 1984, Schippers 2000) may have been responsible for its continual existence and survival. Nevertheless, scientists think that the genetic resources of AYB may have been undergoing gradual erosion.

IITA keeps some accessions of the crop, but otherwise, its conservation in Nigeria is very poor and access to its genetic resources is severely limited. Seeds of AYB seem to be available in the hands of those who appreciate its value, i.e., the elderly farmers and women in a few rural areas in Nigeria. The ancient landraces in the hands of local farmers are the only form of AYB germplasm; no formal hybrid had been produced as yet.

Improvement of the crop is possible only when the intraspecific variability of the large genetic resources of the species is ascertained. The genetic resources of AYB need to be saved for use in genetic improvement through further exploration in tropical Africa and for conservation.

African yam bean plant showing mature pods ready for harvest. Photo by Daniel Adewale, IITA.
African yam bean plant showing mature pods ready for harvest. Photo by Daniel Adewale, IITA.

Understanding AYB
Eighty accessions (half of the total AYB collection under conservation in the IITA genebank) were assessed for diversity using morphological and molecular methods. Thirty selected accessions were further tested in four ecogeographical zones in Nigeria to understand their productivity and stability. The breeding mode was also studied.

Findings show that each of the 80 accessions of AYB has a unique and unmistakable genetic entity, promising to be an invaluable genotype as a parent for crop improvement. Morphologically, two groups have evolved: the tuber forming and the nontuber forming.

Grain yield differed among individual accessions and across the four agroecologies. The average grain yield across the four diverse environments in Nigeria (Ibadan, Ikenne, Mokwa, and Ubiaja) was ~1.1 t/ha; however, grain yield at Ubiaja was well above 2 t. Most agronomic and yield-determining traits had high broad sense heritability and genetic advances, assuring high and reliable genetic improvement in the species. AYB is both self fertilizing and an outcrosser; the latter trait is exhibited at about 10%.

The good news is improvement through hybridization is possible within the species.

References
Anochili, B.C. 1984. Tropical Agricultural Handbook. Pages 48–50 in Food Crop Production. Macmillan Publishers, London, UK.

Bioversity International. 2009. http://www.bioversityinternational.org/scientific_information/themes/neglected_and_underutilized_species/overview.html [25 February 2010].

Devos, P., G.F. Wilson, and E. Delanghe. 1980. Plantain: Genetic resources and potential in Africa. Pages 150–157 in Genetic Resource of Legumes in Africa edited by Doku, E.V. Proceedings of a workshop jointly organized by the Association for the Advancement of Agricultural Science in Africa and IITA, Ibadan, Nigeria, 4–6 January 1978.

Jaenicke, H. and N. Pasiecznik. 2009. Making most of underutilized crops. LEISA Magazine, 25(1):11–12.

Klu, G.Y.P., H.M. Amoatey, D. Bansa, and F.K. Kumaga. 2001. Cultivation and uses of African yam bean (Sphenostylis stenocarpa) in the Volta Region of Ghana. The Journal of Food Technology in Africa 6:74–77.

Naylor, R.L., W.P. Falcon, R.M. Goodman, M.M. Jahn, T. Sengooba, H. Tefera, and R.J. Nelson. 2004. Biotechnology in the developing world: a case for increased investment in orphan crops. Food Policy 29:15–44.

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The cassava scourge

James Legg, j.legg@cgiar.org

Close up of Bemisia tabaci adults. Photo by CIAT
Close up of Bemisia tabaci adults. Photo by CIAT

Who would think that delicate and exquisite little insects such as whiteflies could pose an ongoing and global challenge to humankind’s need to meet its food requirements?

Whiteflies are one of the top 10 most serious pest threats to agriculture. Although whiteflies, in the taxonomic family Aleyrodidae, are a diverse group of insects of more than 1,200 species, only a few of these are economically important. Among this small group, Bemisia tabaci (Genn.) is by far the most important single species.

B. tabaci was first described from tobacco in Greece, towards the end of the 19th century. Its progress has closely matched developments seen in agriculture in subsequent years, and it now occurs virtually throughout the crop-growing parts of the globe. Its preference for warm weather means that it is particularly prevalent in the tropics, although it has also been able to exploit protected agricultural environments in temperate regions.

Deadly partnerships
If B. tabaci contented itself with doing its own thing and sucking small quantities of sap from the plants that it feeds on, it would probably have fallen under the radar of those whose job it is to protect crops. But it did not. Over time, it evolved a relationship with plant viruses, a relationship that allowed the whitefly to pick up viruses when feeding on plants, harbor them for some time, before introducing them to another plant during feeding, thereby giving rise to a new infection. This enabled the viruses transmitted to expand their ranges as B. tabaci populations grew and spread. These deadly partnerships thus gave rise to plant disease epidemics that had devastating impacts on the crops affected, and on the people growing them.

Large population of B. tabaci adults feeding on the underside of a young cassava leaf
Large population of B. tabaci adults feeding on the underside of a young cassava leaf. Photo by IITA

B. tabaci transmits many hundreds of virus species, a number that keep rising as more viruses are described and research efforts on the B. tabaci vector are also broadened. The viruses transmitted fall into four virus genera: Begomovirus (family Geminiviridae), Ipomovirus (Potyviridae), Crinivirus, and Carlavirus (Closteroviridae). More than 90% of the more than 100 species transmitted, however, are in the Begomovirus group. One of Africa’s most economically destructive diseases, cassava mosaic disease (CMD), is caused by a group of viruses in the Begomovirus genus. Collectively, these are usually referred to as the cassava mosaic geminiviruses. Evidence also points to B. tabaci being the vector of cassava’s other major expanding disease threat, cassava brown streak disease (CBSD) caused by the Ipomovirus, cassava brown streak virus.

Cassava has always been at the heart of IITA’s research-for-development agenda. Thus, diseases such as CMD and CBSD, and the agents that promote their spread, have long been the focus of research efforts. From its earliest beginnings, IITA was fortunate to receive cassava germplasm, developed in East Africa through the Amani breeding program that most importantly was endowed with resistance to CMD.

It may have been an unfortunate spin-off of the tremendous success of the CMD-resistant varieties, but B. tabaci, the humble vector of the CMGs, received very little research attention before the 1990s. Things were to change abruptly in the mid-1990s, however. It became increasingly clear that unusually large whitefly populations were propelling the expansion of a new, highly virulent form of CMD in Uganda.

Studying the pandemic
IITA initiated a wide-ranging research program with the dual aims of enhancing scientific understanding of the deadly virus-vector combination as well as working with national partners to manage the pandemic.

The genetics and epidemiology of CMGs associated with the pandemic were extensively documented over the decade following the initial explosion of interest. Although less research attention was focused on the whitefly vector, a number of important advances were made in understanding the nature and role of B. tabaci. Perhaps most significantly, it was demonstrated that superabundance of B. tabaci was a key factor driving the pandemic’s so-called ”front”, and, furthermore, that the front could be pushed forwards by up to 100 km/year in this way. Although whiteflies are weak fliers, single B. tabaci individuals have been shown elsewhere to fly for up to 7 km, assisted by the wind, and given a generation time of slightly less than one month, it is easy to see how such a long distance spread could be achieved.

Superabundant <em srcset=B. tabaci and the CMD pandemic (Yellow shaded area is the approximate region affected by the CMD pandemic by 2009. Arrows indicate the direction of pandemic expansion. White explosions indicate areas in which B. tabaci superabundance has been most prominent, together with associated physical damage to cassava crops.)” width=”250″ height=”181″ />
Superabundant B. tabaci and the CMD pandemic (Yellow shaded area is the approximate region affected by the CMD pandemic by 2009. Arrows indicate the direction of pandemic expansion. White explosions indicate areas in which B. tabaci superabundance has been most prominent, together with associated physical damage to cassava crops.). by IITA

Extensive and regular disease surveys conducted by IITA and its NARS partners from 1997 up to the present have helped build up a comprehensive picture of the pandemic’s expansion into 11 countries of East and Central Africa and the interrelationships with vector populations. Moreover, these data have been used to provide risk assessments of future patterns of spread which have supported disease management initiatives.

Superabundant B. tabaci populations are typically 100-fold greater than those outside the pandemic zone. As well as delivering a sharply increased level of virus transmission, these cause physical damage to cassava plants. Experimental studies conducted at IITA-Uganda showed that yield losses from whitefly damage alone can be as much as 50%, and that these losses are particularly severe for some of the recent releases of CMD-resistant varieties. A gray-black sooty mold covering the lower leaves that develops on the sugary excreta produced by whitefly nymphs is a characteristic symptom of heavy whitefly infestation. These symptoms have been observed in various parts of East and Central Africa, and always occur in areas affected by the CMD pandemic.

Research priority
The obvious research question that has been thrown up from these sets of circumstances is: ”what causes superabundance in B. tabaci?” There are two principal hypotheses. One suggests that superabundance is a result of the spread of a novel ‘fitter’ B. tabaci biotype, and the second, that superabundance is the consequence of a synergistic interaction between B. tabaci and CMD-infected cassava plants.

To examine the first hypothesis, IITA has been working with the University of Arizona, USA, to develop molecular markers to allow discrimination between cassava-colonizing B. tabaci populations. The earliest work made use of sequence portions of the cytochrome oxidase 1 gene of mitochondrial DNA (mtCO1). MtC01 sequences were obtained from whiteflies collected along transects straddling the pandemic ”front” in Uganda. Analysis of sequence homologies showed that there were two major genotype clusters, and that one of these, the so-called “invader” was strongly associated with the pandemic-affected zone. Subsequent collections made after the pandemic had covered the whole of the cassava-growing area of Uganda, however, provided an apparently contradictory outcome, as the ”invader” genotype cluster appeared only infrequently. This is not altogether surprising, however, as B. tabaci cassava biotypes from different countries, and even different continents, have been shown to be able to interbreed successfully.

Finding novel solutions
Current efforts are therefore focusing on developing microsatellite markers that provide a much wider coverage of the B. tabaci genome and will make it more likely that we can discriminate between putative superabundant and normal B. tabaci biotypes. To investigate the intrinsic biological characteristics of different cassava B. tabaci populations, their associated genetics and the biology of offspring produced through inter- and intra-population mating, core funds are currently being used to run a PhD program in Tanzania. This study will also be used to examine the hypothesis of B. tabaci-CMD infected cassava synergism. Preliminary results from cage trials conducted at NRI using a single variety have shown increased B. tabaci abundance on CMD-infected plants, when compared with uninfected material.

Chlorosis on shoot tip and sooty mold on lower leaves caused by heavy <em srcset=B. tabaci infestation” width=”250″ height=”188″ />
Chlorosis on shoot tip and sooty mold on lower leaves caused by heavy B. tabaci infestation. Photo by IITA

The idea that diseased cassava makes for a better food source for B. tabaci has parallels in studies conducted with B. tabaci on other host plants, where virus infection has led to increased whitefly populations. In the cassava system there are some contradictions, however. It is significant that the greatest abundances of B. tabaci in pandemic-affected areas are actually observed on CMD-free resistant varieties. Further research is clearly required before a clear-cut explanation can be given for the superabundance enigma.

With whitefly numbers at record levels, and physical damage exacerbating the already grave problems posed by CMD, it has been increasingly recognized that effective measures for whitefly control need to be identified. Two main options appeared to offer greatest potential: resistance and biocontrol. Pesticides, although widely used in northern commercial agricultural systems, are easily dismissed for use on cassava in SSA, because of the extreme cost and the environmental hazard that they pose.

Is biocontrol the answer?
IITA had great success in its classical biological control programs for managing cassava mealybug and cassava green mite. Why not do a similar thing for whiteflies? Sadly, B. tabaci poses a greater challenge since it is considered to be African in origin, and therefore should already be benefiting from the presence of indigenous natural enemies. Significant work was nevertheless undertaken at IITA-Uganda to characterize the natural enemies of B. tabaci on cassava and to investigate the potential for augmentation.

A combination of surveys, life table studies, mortality measurements, and behavioral assessments conducted over a 10-year period—from 1999 to 2008—revealed that although natural enemies accounted for significant mortality in B. tabaci populations, under normal circumstances this was not sufficient to keep B. tabaci populations at levels below those causing significant economic damage.

To change this balance, it was concluded that complementary B. tabaci control measures would be required, such as the introduction of climate-matched exotic B. tabaci parasitoids or the use of cassava varieties either less favorable to whiteflies or more favorable to parasitoids. Although no attempt has yet been made to introduce exotic B. tabaci parasitoids to East Africa, a significant amount of effort has been made to enhance whitefly resistance in cassava germplasm. IITA partnered with CIAT, NRI, and NARO (Uganda), under the SP-IPM’s Tropical Whitefly Project, to pioneer efforts to introduce to East Africa strong sources of whitefly resistance developed in Latin America by CIAT (albeit to different whitefly species).

The NARO team have had some success in identifying Latin American germplasm that appears to have partial resistance to African B. tabaci, but the challenge still remains to combine these sources of resistance with the other key traits that are required by cassava in the East African farming environment. To achieve this, whitefly resistance markers will need to be built in to marker-assisted selection approaches. Much untapped potential may yet exist, however, in African germplasm, and beyond that, within wild relatives. These are important areas of future research.

Women bringing cassava to market
Women bringing cassava to market. Photo by IITA

Need more studies on whitefly
The recent upsurge in the importance of CBSV in the Great Lakes region of East/Central Africa poses yet more challenges to the cassava research community. Although published reports identify B. tabaci as the vector, researchers remain divided on the accuracy of this claim. As such, IITA, working closely with NRI, is actively addressing this question systematically, by combining field epidemiological studies with cage-based transmission experiments, both of which are being facilitated by newly improved virus diagnostic techniques.

Preliminary results seem to support the earlier claim that B. tabaci is the vector, as the level of CBSV infection in whitefly-protected experimental plots was approximately half that in whitefly-infested plots. These preliminary data will need to be confirmed by repeat trial plantings and cage trial results before any more definitive outcome can be claimed.

Whiteflies have been recognized as an important threat to cassava production for more than a century, but at the outset of the 21st century, that threat appears to be greater than ever. It appears likely that B. tabaci is driving a dual pandemic of CMD and CBSD through the cassava-growing heartlands of Africa.

Recognition of the importance of the twin threats to cassava is at an all-time high, with record levels of funding available to tackle them. By contrast, the role of the vector in the cassava crisis has received much less recognition. This fact will need to be addressed by IITA and its partners in developing future cassava-oriented R4D projects and programs.

Developing genomic resources for banana

Jim Lorenzen, j.lorenzen@cgiar.org

jim_lorenzen-looking-at-banana-flower
Jim Lorenzen checking a banana flower. Photo by IITA

Banana and plantain (Musa sp.) are a very important staple food and cash crop in Africa. Although the principles of banana breeding and genetics were established decades ago, it is still a time-, land-, and resource-intensive process. A crew of several persons collects male flowers and pollinates female flowers while perched on ladders. When successful, seeds must be surface-sterilized and embryos removed for germination in test tubes (or else most won’t germinate), multiplied, and carefully “weaned” for field planting.

Large-sized plants require much field space, and new hybrids must be evaluated through two or three production cycles (about 3 years) before being selected for further testing, such as for disease resistance. Some essential attributes, such as resistance to disease or nematodes may also take several years to assess properly. It would be a huge advantage if early selection could be done, based on some associated marker or rapid test, to eliminate susceptible individuals without wasting resources on them. For other complex traits, it would be useful to have markers based on component genes to be able to select ideal “genotypes”.

One way to do early selection is to use molecular markers that are linked to the target traits (molecular-assisted breeding). This method is becoming common in cereal breeding, yet should be even more cost-effective for a large perennial crop such as banana that requires so much time and space to evaluate. The tools of DNA fingerprinting are applied, and by knowing which DNA markers lie near genes of interest, selecting for the markers will be equivalent to selecting for the trait a year or more later.

The problem is that we lack enough information on the banana genome to have molecular tools to map traits. One of our activities has been to map and characterize new molecular markers for use in banana breeding and genetics. PhD student Gaby Mbanjo from the University of Yaoundé, Cameroon, has been working in Uganda and Kenya to characterize and map a large new set of simple sequence repeat (SSR) markers, often called microsatellite markers. She is a scholar of the Biosciences for Eastern and Central Africa (BecA) program, with funding provided by the Canadian International Development Agency (CIDA).

Gaby is also working to develop other types of molecular markers based on small genetic differences (single nucleotide polymorphisms = SNPs) between alleles of genes involved in controlling plant defensive reactions. These will be used to try to map the genetic loci responsible for resistance to the burrowing nematode (Radopholus similis) and banana weevil (Cosmopolites sordidus) in the population she is studying. Markers of both types can be converted to semi-automated assays for hundreds or thousands of assays. This effort is expected to result in a DNA fingerprinting assay in which we can select the associated DNA markers and thus also select the target resistance without spending as many resources on susceptible plants.

The molecular markers will have other practical uses. Unfortunately, sometimes varieties get distributed with wrong names, or a batch of plants supposedly of a single variety may actually contain a mixture of varieties. Molecular markers can be used to “fingerprint” mother plants used to produce new planting material to ensure that they are of the proper variety. They can also be used to select diverse parents for maximizing the heterozygosity of offspring. Some of the markers are being transferred to a national research program for assessing varietal purity in their advanced selections.

Molecular markers are a way in which biotechnology and the rapidly expanding knowledge of DNA sequences in plant genomes can be used to make classical breeding more efficient. This should be especially helpful for large perennial crops such as banana and plantain.

Biotechnology and nematodes

Leena Tripathi, l.tripathi@cgiar.org

Banana and plantain (Musa spp.) are major staple foods and a source of income for millions in tropical and subtropical regions. Most of the banana grown worldwide are produced by small-scale farmers for home consumption or sale in local and regional markets.

Leena Tripathi inspecting a diseased banana leaf. Photo by IITA
Leena Tripathi inspecting a diseased banana leaf. Please note that this picture does not relate to nematode damage or diseased by nematodes as correctly pointed out by Danny Coyne, IITA, thanks!Photo by IITA

Many pests and diseases significantly affect banana cultivation. Nematodes pose severe production constraints, with losses estimated at about 20% worldwide. Locally, however, losses of 40% or more occur frequently, particularly in areas prone to tropical storms that topple the plants.

Pest management in banana is based on several principles, primarily through the use of clean, healthy planting material, crop rotation, and in commercial settings, chemical control. However, crop rotation is often impractical, especially for small-scale farmers, while nematicides are locally unavailable or not affordable for subsistence farmers. These pesticides are also highly toxic, environmentally unacceptable, and increasingly being withdrawn from use.

Limited sources of nematode resistance and tolerance are present in the banana gene pool. Some resistance has been identified against one of the most damaging nematode species, the burrowing nematode (Radopholus similis), but this needs to be combined with consumer-acceptable traits. Furthermore, several species of nematodes are often present together, requiring a broad spectrum resistance able to control not just Radopholus but other damaging nematodes, such as species of Pratylenchus, Meloidogyne, and Helicotylenchus.

Enter biotechnology. Biotechnology offers sustainable solutions to the problem of controlling plant parasitic nematodes. Several approaches are possible for developing transgenic plants with improved resistance; these include strategies against invasion and migration and against nematode feeding and development.

Woman selling banana in a local market. Photo by IITA
Woman selling banana in a local market. Photo by IITA

Some successes in genetic engineering of banana have been achieved, enabling the transfer of foreign genes into the plant cells. An efficient transformation protocol for African banana cultivars has been established at IITA using meristematic tissues. The protocol avoids the callus and cell suspension culture requirements of other approaches. It is rapid, genotype independent, and avoids the somaclonal variation that often results from regenerating embryogenic cell suspensions.

IITA, in partnership with the University of Leeds, UK, is exploring the potential of biotechnology to develop plantain resistant to nematodes with funds from the Department for International Development/ Biotechnology and Biological Sciences Research Council.

Prof. Howard Atkinson’s group in Leeds has demonstrated that more than one independent basis for transgenic resistance provides an additive effect for nematode control. Our use of three independent additive approaches is designed to ensure a resistance level that prevents the buildup of damaging populations, even if virulent individuals completely challenge one line of defense or partially compromise them all. We intend to demonstrate that this additive approach can provide durable resistance.

nematodes
nematodes, photo by IITA

The three approaches are a cysteine proteinase inhibitor (a cystatin), a potato tuber serine/aspartic proteinase inhibitor, and a repellent peptide. Cysteine proteinases are used by a wide range of plant parasitic nematodes to digest dietary protein. The cystatin prevents this digestion and slows nematode growth. Transgenic expression of both proteinase inhibitors provides effective control of both cyst and root-knot nematodes and cystatin has also been shown to be effective against Radopholus.

Cysteine proteinases are not present in mammals and those we will use lack toxicity or allergenicity for humans. They occur in common foods, such as the seeds of maize, rice, and cowpea, and people rapidly digest them. The other, very distinct, novel approach is the use of a repellent. This is also not lethal to nematodes or other organisms. Nematodes do not invade roots applied with repellent because they fail to detect the host’s presence. This approach is effective against a wide range of nematode species.

We will also be using a novel RNA interference (RNAi) approach. The use of RNAi for functional analysis of plant parasitic nematode genes was first established in the University of Leeds. The approach relies on the production of double-stranded RNA molecules by banana cells. When they are ingested by the nematode, they specifically interfere with the expression of the essential nematode gene they target. The advantage of the RNAi approach is that no novel protein production is required to achieve resistance to nematodes. This offers a considerable biosafety advantage, given that RNA molecules represent no food risk and there is little likelihood of nontarget effects. The challenge is to provide an effective level of resistance to all banana nematodes by this approach. Genetic transformation of plantain using these approaches is in progress at IITA.

Banana field trials in Rwanda. Photo by IITA
Banana field trials in Rwanda. Photo by IITA

Gene flow is not an issue for this crop, making the transgenic approach even more attractive. Banana and plantain lack cross-fertile wild relatives in many production areas. Most edible banana are male- and female-sterile and depend on vegetative propagation. The new defense will be integrated with other pest management strategies already developed at IITA to maximize resistance levels and safeguard durability. This work is part of a new, interdisciplinary research partnership between IITA and the University of Leeds, directed at enhancing human health and food security in sub-Saharan Africa.

We also plan to stack genes for Xanthomonas wilt and nematode resistance into one line to produce a high-value product for farmers. Gene stacking is becoming common, adding multiple traits at once into the plant genome. Resistance to diseases and pests can be achieved by integrating several genes with different targets or modes of action into the plant genomes. We already have promising results with genes for resistance to banana Xanthomonas wilt (BXW) (R4D Review Edition 1), which we would like to combine with nematode resistance. Banana cultivars with resistance to multiple diseases and pests will be a breakthrough in banana improvement.

Designer (cowpea) plants

Christian Fatokun, c.fatokun@cgiar.org

Plants can be designed to order. Science has long found a way to combine good and useful characteristics in a plant by studying the genes for such traits, and putting them together in a process called “genetic engineering.”

Caterpillar boring into a cowpea pod. Photo by S. Muranaka
Caterpillar boring into a cowpea pod. Photo by S. Muranaka, IITA

Cowpea is grown mainly for its protein-rich grains and quality fodder for livestock. At present, biological control and conventional breeding methods are proving inadequate in developing cowpea varieties resistant to destructive pests, such as the legume pod borer Maruca vitrata.

M. vitrata is the most widespread cowpea pest. The adult moth lays eggs on the plant. The larvae that emerge from the eggs damage plants in the field, particularly during the reproductive stage, through feeding on young succulent shoots, flowers, pods, and seeds. This pest can cause significant grain yield reduction, between 20% and 80% if not controlled with insecticides.

Farmers usually spray insecticides to protect the cowpea crop from Maruca and other pests. Purchasing chemicals, however, adds to the production cost, thus reducing the farmers’ profit. Also, farmers are not well equipped to protect themselves when using such toxic chemicals. In some farming communities, adulterated chemicals that do not control the pests are sold to farmers. The development of cowpea varieties with resistance to Maruca and other insect pests would benefit the most resource-poor African farmers who grow the crop.

Cowpea is grown extensively in the savanna region of sub-Saharan Africa (SSA). At least one major insect pest attacks cowpea at every stage in the life cycle, including seeds in storage. These pests are significantly responsible for the low grain yield in farmers’ fields.

Through conventional breeding, some varieties have been developed that show resistance to some of the pests, such as aphids and flower thrips, and low levels of resistance to the storage weevil. However, not much progress has been made in host plant resistance, especially M. vitrata.

Efforts continue to identify parasites and predators that could be used as biocontrol agents. When deployed, such agents would greatly reduce the population of the Maruca larvae in the field, giving the cowpea plant some respite for the production of flowers and pods containing whole and well-formed seeds.

Using conventional breeding, several hundreds of accessions of cultivated cowpea and its wild relatives have also been screened for resistance to this pest. Accessions belonging to Vigna vexillata were found to be resistant to M. vitrata. These accessions were found to be closest to cowpea in a phylogenetic study of diversity in the Vigna species. The study was based on data obtained after DNA genotyping. Efforts were made to cross cowpea with V. vexillata but without success.

This strong cross-incompatibility makes gene exchange between the two species impossible. This is where biotechnology comes to the rescue. Two major steps are needed to develop genetically modified cowpea with resistance to M. vitrata. First is developing a transformation system and the second is identifying the transgene that would be effective against the pest when introduced into cowpea. Since Maruca is a Lepidopteran, some of the genes from Bacillus thuringiensis (Bt) should be effective against the insect’s larvae. IITA screened several Bt protoxins on Maruca by incorporating different concentrations in the diet fed to the larvae. The protoxin of Bt gene ”Cry1ab” was found to be most effective even at very low concentrations in the artificial diet. This Bt gene (Cry1ab) was therefore selected as the candidate gene for designing Maruca-resistant cowpea.

Scientists at the Commonwealth Scientific and Industrial Research Organisation (CSIRO) in Australia developed the transformation system using an IITA-developed breeding line ”IT86D-1010” derived from a cross between ”TVx4659-03E” and ”IT82E-60”. CSIRO scientists have now obtained cowpea lines containing the Bt gene. Monsanto donated the Bt gene used to transform cowpea by licensing it to the African Agricultural Technology Foundation (AATF) for use in Africa. The Rockefeller Foundation and USAID funded this cowpea transformation project.

Farmers transporting cowpea harvest. Photo by S. Muranaka
Farmers transporting cowpea harvest. Photo by S. Muranaka, IITA

The Bt cowpea had been tested in the CSIRO laboratory in Australia and found to be effective against the larvae of another Lepidoptera, Helicoverpa armigera. The Bt gene in cowpea is expected to be effective against M. vitrata, the cowpea pest, but needs to be tested in an environment where Maruca thrives. Apart from Burkina Faso, none of the African countries where cowpea is an important crop has a biosafety law in place. A few lines of the Bt cowpea were, therefore, taken to Puerto Rico for field testing. The field trial was carried out in late 2008. If the Bt gene in the cowpea lines is found to be effective against Maruca, the next step would be to transfer the Bt gene into popularly grown cowpea varieties selected from interested countries. The line presently containing the Bt gene is not high yielding, and farmers are not likely to accept it readily.

Under the international biosafety protocol (Cartagena protocol on biosafety) it is necessary to carry out risk assessment on the Bt cowpea before it is introduced to another country. The data obtained from risk assessment form part of the dossier that accompanies applications requesting for importation to any country. Risk assessment would entail studies on gene flow, the effect of the transgene on nontarget organisms, food safety, and resistance management strategies.

A meeting of experts in these various fields is planned in March 2009 at the Donald Danforth Plant Science Center, St. Louis, Missouri, USA. The experts would design studies to address the different questions that may arise from biosafety regulators in the countries where the Bt cowpea is meant to be grown. Many of the proposed studies are necessary, because cowpea is an indigenous food crop in SSA where cross-compatible wild relatives are found growing in agroecologies similar to farmers’ fields. Biosafety reviews in the African countries would, therefore, be rigorous.

Organic bananas from Africa?

Organic agriculture provides significant market opportunities for commercial agriculture globally.

Organic food markets grew at tremendous rates during the 1990s, encouraging organic food production throughout the world. Although this growth rate has slowed down a bit, and the niche market for organic food is less than 4% of the European or North American food markets, the prospects of high prices and a stable demand still make organic food markets attractive for producers.

Commercial and certified organic farming is not uncommon in Africa that has 19% of the world’s organic farms. Main organic products include fruits and vegetables, cotton, coffee, tea, and herbs and spices.

Bananas are the most widely traded fruits worldwide. Recent trends in organic food demand in developed countries have made organic bananas an attractive crop in developing countries. In fact, trade in organic bananas increased during the late 1990s and early 2000s at a quite significant rate, from about 30,000 tons in 1998 to about 150,000 tons in 2003. Even so, organic bananas represent only a small share (1%) in the world banana trade.

About two-thirds of the organic bananas are traded to the European Union (EU), where they constitute about 2.5% of the banana market, a significantly larger percentage than on the world market. The other main target is North America, and, to a much lesser extent, Japan.

The entire trade in organic bananas comes from countries in Latin America. Africa and Asia are geographically closer to the EU and Japan but this does not seem to be relevant to organic banana trade.

Conventionally grown bananas are mainly traded from Latin America onto the world market, with minor shares coming from West and Central Africa (WCA) and Asia. North America sources its banana supply exclusively from Latin America. Europe imports bananas from Latin America, WCA (Côte d’Ivoire and Cameroon). Japan imports bananas from Latin America and Asia (Philippines and China).

Market stall in Tanzania showing range of local banana types. Photo by IITA
Market stall in Tanzania showing range of local banana types. Photo by IITA

East and Central Africa—in particular Uganda, Rwanda, and Burundi, as one of the largest banana-producing regions worldwide—does not feature to any significant extent in these statistics.

Suppliers of organic bananas are basically the same as those of conventional bananas, Latin American countries. Organic banana production and trade follow conventional production and trade, with suppliers such as the African or Asian producers lagging behind.

Although this is a large market, bananas, in particular conventionally grown bananas, seem to have had the peak of their market growth during the 1990s. Significant volume growth is expected to occur only in Eastern Europe and the Middle East. Elsewhere, volume growths are expected only to follow population developments, to a lesser extent increases in income and falling prices. While volume growths reached on average 4% in the 90s, they will reach only about 2.5% annually until 2010. Prices are expected to decline with increased liberalization of banana markets, in particular the EU. Overall, markets are considered saturated.

Africa has not been able to take up production and trade opportunities on the global banana market, with a few exceptions such as Cameroon and Côte d’Ivoire. However, more recently, there are efforts under way to try and enter the global banana markets in both the conventional and niche segments.

Conventionally grown banana production in Uganda, Rwanda, and Burundi is hardly competitive with that from other regions because of its small scale and low-input production. These lead to relatively low yields and consequently high production costs. Scattered small-scale production makes assembly and packaging a long and costly effort, with high postharvest losses as a consequence.

Transport routes are long and road and sea transport to possible final destinations often take longer than the shelf-life of bananas, so that the freight will decay before reaching markets in Europe or Asia.

Consequently, the only exports of conventionally grown bananas from East Africa to Europe go by airfreight, often as by-cargo with higher value products. In Europe they supply only specialty markets, such as cooking bananas or plantain for African expatriates, who do not make up a significant market share. The problem can be quantified by comparing production and trade costs of conventionally grown bananas from Latin America and Uganda (Table 1).

The reasons for Uganda’s disadvantages on the European market are obvious: High raw material costs. Land and labor-intensive small-scale production, losses from pests and diseases, and the lack of fertilizer already affect primary production adversely. Gathering, packaging, and transport from the small farms through many intermediaries impose a large amount of additional costs.

The transport of the material to the seaports (the nearest is Mombasa in Kenya) and the long distance to Europe add further disadvantages. Normally, the distance from production to market in terms of days would exceed even the 20-day shelf-life of fresh bananas.

Organic bananas from Uganda are cheaper at the farmgate than Ecuadorian bananas (Table 1), and although handling costs and airfreight are still high, and the final margins in retail are lower than those from Ecuadorian bananas, there is still a significant profit margin at the retail level. This makes the export of organic bananas from Uganda to the EU by airfreight far more attractive than the export of conventional bananas by sea—if the latter becomes technically possible.

This opportunity should be the same for more Central African countries, such as Rwanda and Burundi, but also for West African countries such as Cameroon and Côte d’Ivoire. What mainly contributes to this opportunity is the high value of organic bananas on the European markets, and the opportunities arising from this to export these high-value fruits to Europe by plane.

However, even if organic bananas (or any other organic fruit or agricultural product) represent an opportunity, some challenges exist which have to be considered. Poor quality and badly maintained roads, vehicles, rail links, and rolling stock all pose problems for transportation. Lack of refrigeration, erratic power supplies, poor communications, underdeveloped banking and credit systems, and, sometimes, political and economic instability, all raise serious and often insuperable problems.

In addition, the lack of local certification bodies imposes significant constraints and risks to organic agriculture in Africa. Certifiers have to be flown in and they increase the costs of organic production. So far, only Tunisia has its own European-standard certification bodies. The costs of certification have to be seen as investment costs and hence risks. If the investment costs are not amortized by the revenues, e.g., in the case of harvest failures or a sudden shortfall of market outlets, investments in certification are lost and hence, the farmers are liable to a significant investment risk. Similar constraints apply to the establishment and sustainability of commercial organic agriculture elsewhere in Africa, and also to the production and trade in organic bananas.

In summary, these constraints are:
•    Lack of experience in intensive organic production
•    Lack of experience in handling and exporting fresh produce
•    Lack of professional management
•    Diseconomies of scale in exporting small quantities, e.g., for test exports
•    Poor communication between foreign importers and exporters
•    Poor negotiation skills and judgment of exporters
•    Lack of familiarity with international markets, including knowledge of the organic market place overseas
•    Lack of governmental support for exports

Selling bananas in the local market. Photo by IITA

Organic banana production has its advantages, in particular for some Central and Eastern African producers, as markets are high value and stable in Europe and the US, while conventional banana markets are stagnating. Yet it is clear that there are a number of prerequisites for entry to the markets. Good marketing linkages and marketing skills for producers and marketers are at the top. Investments in certification have to be facilitated, in particular for small producers or producer groups.

Both physical infrastructure (roads) and political frameworks in Africa have to be favorable if organic production and exports are to be sustainable. Markets, although attractive at the moment, are competitive, probably limited, and probably highly income-elastic and thus sensitive to economic distortions on the demand side. This also means that oversupply has to be avoided and in the long run, cost reduction will be necessary to successfully compete in organic markets.

Sources:
FAO. 2004. Statistical databases. www.fao.org.
Fischer, C. 2004. Demand for bananas in the European Union, with special focus on Germany. Research report, Bonn.
Mwadime, S. 2004. Private sector developments. Paper presented on the conference “Markets to raise incomes for poor farmers in Africa”, organized by the Rockefeller Foundation in Nairobi, Kenya, 5-8 April 2004.
Spilsbury, J., Jagwe, J., Ferris, S. and D. Luwandagga. 2002. Evaluating the market opportunities for banana and its products in the principal banana growing countries of ASARECA, Uganda report. Kampala (IITA/Foodnet).
FAO. 2006. Medium term prospects for agricultural commodities: Bananas. http://www.fao.org/docrep/006/y5143e/y5143e10.htm.
FAO/ITC/CTA. 2001. World Markets for Organic Fruit and Vegetables – Opportunities for Developing Countries in the Production and Export of Organic Horticultural Products. International Trade Centre, Technical Centre For Agricultural and Rural Cooperation, Food and Agriculture Organization of The United Nations, Rome, Italy. Online at http://www.fao.org/docrep/004/y1669e/y1669e00.htm.
Parrott, N. and F. Kalibwani. 2005. “Organic Farming in Africa,” In The World of Organic Agriculture: Statistics and Emerging Trends 2005. Helga and Yussefi (Eds). International Federation of Organic Agriculture Movements (IFOAM), Bonn, Germany and Research Institute of Organic Agriculture (FiBL), Frick, Switzerland.
Parrot, N. and van Elzakker, B. 2003. Organic and like-minded movements in Africa. Development and status.- IFOAM.
UNCTAD (2008): Market information in the commodities area: Banana. http://www.unctad.org/infocomm/anglais/banana/market.htm.
Yussefi, M. 2006. “Organic Farming Worldwide 2006: Overview and Main Statistics,” In The World of Organic Agriculture: Statistics and Emerging Trends 2006. 7th Revised Edition, Willer and Yussefi (Eds). International Federation of Organic Agriculture Movements (IFOAM), Bonn, Germany and Research Institute of Organic Agriculture (FiBL), Frick, Switzerland.

The future of African bananas

The use of genetic engineering has transformed agriculture, and food production and development by providing options and solutions where none existed before—to the benefit of billions of the world’s inhabitants.

Tripathi discusses work with staff at Namuloge, IITA-Uganda. Photo by IITA

IITA and its partners have been using genetic transformation as a crop improvement tool to help produce more and better food staples. The Institute—with partners such as the National Agricultural Research Organization (NARO) of Uganda, Academia Sinica (Taiwan), and the African Agricultural Technology Foundation (AATF) in Kenya—is at the forefront of research “designing” a genetically modified banana that is resistant to the worst bacterial disease so far—Banana Xanthomonas Wilt (BXW). Entire banana fields can be destroyed, especially those planted to Pisang awak, a susceptible exotic variety widely grown to make banana beer.

Bananas are a major staple in East Africa produced mostly by smallholder subsistence farmers. Uganda is the world’s second leading grower with a total annual production of about 10.5 million tons. It is Africa’s biggest producer and consumer of bananas and plantains.

Most growers cannot afford costly chemicals to control the many pests and diseases that affect banana cultivation. As diseases continue to spread, demand grows for new improved varieties.

Bacterial wilt caused by Xanthomonas campestris pv. musacearum is threatening banana production and the livelihoods of these smallholder growers, and solutions have to be found fast before it could destabilize food security in the region.

Work on developing a GMO banana has been ongoing since BXW was first reported in 2001. The disease has been identified in the Eastern Democratic Republic of Congo, Rwanda, Kenya, and Tanzania, and is widespread in Uganda. It attacks almost all varieties of bananas, causing these countries an annual loss of over US$500 million. These can be reduced bunch weights or absolute yield loss or clean planting material is unobtainable for new plantations.

Banana Xanthomonas wilt-infected plants. Photo by IITA

“Developing resistant varieties is a long-term but more sustainable way to control pests and diseases. Improving the plant’s defense mechanism against BXW through genetic engineering is still the best line of defense because of its many advantages,” commented molecular geneticist Leena Tripathi based in IITA-Uganda, Kampala. “Farmers are reluctant to employ labor-intensive disease control measures.”

“Genetic engineering offers many opportunities for improving existing elite varieties not amenable to conventional cross-breeding, such as bananas. It allows breeders to develop new varieties quickly through the introduction of cloned genes into commercial varieties.”

Transgenic bananas possess a gene or genes that have been transferred from another plant species. The term “transgenic plants” refers to plants created in a laboratory using recombinant DNA technology.
Tripathi said that the development of stable and reproducible transformation and regeneration technologies has opened new horizons in banana and plantain breeding. The development of transgenic banana and plantain has been reported by several groups, but a commercial transgenic banana variety is yet to be released.

There are no cross-fertile wild relatives in many banana-producing areas. Most edible bananas and plantains are male and female sterile. The clonal mode of propagation makes the risk of gene flow from banana to another crop species not an issue.

IITA’s in vitro screening method for early evaluation of resistance to BXW uses small tissue culture-grown plantlets. This method can be used by breeders for screening Musa germplasm with larger numbers of cultivars for resistance to BXW and other bacterial diseases.

Currently, most transformation protocols for banana use cell suspensions, Tripathi said. Establishing cell suspensions is a lengthy process and cultivar dependent. At present, the major barrier in transforming East African Highland Bananas (EAHB), a cooking banana from Uganda, is the limited success in producing embryogenic cell suspension cultures from a wide range of cultivars. IITA scientists in collaboration with NARO have developed a rapid and efficient protocol using a cultivar-independent transformation system for improving Musa species including EAHB. This new technique has paved the way for the development of a transgenic banana using transgenes from sweet pepper that confer resistance against BXW.

Tripathi explains how the technology works: the ferredoxin-like amphipathic protein (pflp) and hypersensitive response-assisting protein (hrap), were isolated from the sweet pepper, Capsicum annuum. These are novel plant proteins that intensify the harpinPSS-mediated hypersensitive response (HR). These proteins have a dual function: iron depletion antibiotic action and harpin-triggered HR enhancement. The transgenes were shown to delay the hypersensitive response induced by various pathogens in nonhost plants through the release of the proteinaceous elicitor, harpinPss in various crops including dicots such as tobacco, potato, tomato, broccoli, orchids and monocots such as rice. Elicitor-induced resistance is not specific against particular pathogens, hence it is a very useful strategy.

The pflp genes encode for ferrodoxin, which exists in all organisms, and is therefore common in human diets. This protein is safe for human consumption and the environment. The pflp and hrap genes are owned by Taiwan’s Academia Sinica, the patent holder. IITA has negotiated a royalty free license through the AATF for access to the pflp and hrap genes for use in the production of BXW-resistant varieties in sub-Saharan Africa.

Hundreds of transformed lines of various banana cultivars have already been generated, and are under screening for disease resistance under laboratory conditions. The most promising will be evaluated for efficacy against BXW in confined field trials under different farming systems by national partners with IITA. The transgenic lines will be tested for environmental and food safety, in compliance with target country biosafety regulations, risk assessment and management, and seed registration and release procedures. The project will also study public perceptions, consumer preferences, and the acceptability of transgenic banana in Africa to guide commercialization and wide use.

“Wide-scale deployment of genetically modified, farmer-preferred banana varieties in African countries would succeed only with effective interinstitutional partnerships, particularly with advanced research institutions, AATF, national committees on biosafety, nongovernmental organizations, and private tissue culture companies,” explained Tripathi. “This project will enhance the capacity of partners from the national agricultural research and extension systems in genetic transformation of banana, molecular biology, and biosafety. High-yielding BXW-resistant banana will bring greater productivity for smallholder farmers in East Africa and improved food security.”