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Has the introduction of modern rice varieties changed rice genetic diversity in a high-altitude region of Nepal?
Field Crops Research 113 (2009) 24–30
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Has the introduction of modern rice varieties changed rice genetic diversity in a high-altitude region of Nepal? K.A. Steele a,*, S. Gyawali b, K.D. Joshi a, P. Shrestha b, B.R. Sthapit c, J.R. Witcombe a a
CAZS Natural Resources, Bangor University, Gwynedd LL57 2UW, UK LI-BIRD, PO Box 324, Giarapatan, Pokhara, Nepal c Bioversity International, Office for South Asia, National Agricultural Science Centre, DPS Marg, Pusa Campus, New Delhi 110012, India b
A R T I C L E I N F O
A B S T R A C T
Article history: Received 12 December 2008 Received in revised form 1 April 2009 Accepted 4 April 2009
We surveyed the uptake of three modern rice varieties by farmers in high-altitude villages in the Kaski district of Nepal and found that their uptake had displaced some traditional landraces in the district. The three varieties, Machhapuchhre-3 (M-3), Machhapuchhre-9 (M-9) and Lumle 2, were developed using client-oriented, participatory plant breeding methods and the first was introduced to farmers in 1996. By 2004 up to 60% of the land area was used to grow these modern varieties. Molecular markers (SSR) were used to assay levels of genetic diversity to test if adoption of modern varieties in the place of landraces had changed genetic diversity. The modern varieties were found to contain diverse alleles with a high proportion from the local parent variety, Chhomrong Dhan. We found a high level of allelic richness in the landraces, and although seven had been dropped in favour of the modern varieties, other diverse landraces were still being cultivated by farmers in the study villages on up to 40% of the rice area. Genetic diversity may be maintained even when landraces are displaced by modern varieties. Using a model we found that the partial replacement of landraces increased genetic diversity if the modern varieties were adopted on up to 65% of the area. Only above these levels did overall diversity decline. ß 2009 Elsevier B.V. All rights reserved.
Keywords: Client oriented breeding (COB) Participatory plant breeding (PPB) Rice Genetic diversity Genetic erosion Traditional variety Landrace Nepal
1. Introduction The adoption of modern varieties by farmers is a key development factor in Nepal that is starting to change cultivation practices and may promote genetic erosion (Bardsley and Thomas, 2005). In areas with harsh environments to which landraces are adapted genetic erosion could lead to the loss of important adaptive traits. An example is cold-tolerance in high altitude (1300–2200 m) rice landraces. The high-altitude rice area in Nepal is about 2% of the total 1.5 m ha rice growing area of Nepal, and until the mid-nineties very few modern varieties had been bred for these domains. Here cold injury due to cool air temperature is a major constraint to improving rice productivity (Sthapit, 1992) and plants are exposed to cold irrigation water (16–22 8C) during the reproductive phase. Three modern varieties for high altitudes of Nepal were developed in Kaski district through the successful use of clientoriented, participatory methods. Participatory plant breeding describes the activity (Morris and Bellon, 2004), whereas client oriented breeding (COB) describes its purpose (Witcombe et al.,
* Corresponding author. Tel.: +44 1248 388655; fax: +44 1248 370594. E-mail address: [email protected] (K.A. Steele). 0378-4290/$ – see front matter ß 2009 Elsevier B.V. All rights reserved. doi:10.1016/j.fcr.2009.04.002
2005). This breeding programme led to the release of Machhapuchhre-3 (M-3) in 1997 (Sthapit et al., 1996; Joshi et al., 1997). Subsequently a sister line Machhapuchhre-9 (M-9) was identified as being highly desirable (Joshi et al., 2001). In 2001, a third modern variety, Lumle 2, from a different cross was identified as suitable by farmers in high altitude villages in Kaski. Although not yet formally released, both M-9 and Lumle 2 have been adopted by farmers and are spreading through farmer-to-farmer seed networks. The effects of introducing modern varieties on changes in the cultivation of rice landraces between 1996 and 1999 in highaltitude regions of Kaski were recorded by Joshi et al. (2001) and Joshi and Witcombe (2003). Landrace cultivation decreased as a proportion of the total area under cultivation and by fewer households growing a given landrace, however few landraces were dropped completely. The adoption of modern varieties caused a greater decline in the more commonly grown landraces while the less common ones were not replaced so rapidly because farmers kept them for their specific adaptations and uses. Participatory breeding methods are intended to produce better quality and higher yielding varieties that are adapted to local cultivation, however there are widespread concerns that the uptake of modern varieties may have negative impacts on genetic diversity (Brush, 2000). Witcombe (1999) has argued that such
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breeding methods do not necessarily lead to a loss of genetic diversity, and that use of participatory methods may even widen crop genetic diversity, particularly where existing biodiversity in farmers’ fields is low. We hypothesised that by adopting modern varieties bred from a cross between a local variety and an exotic variety, the total allelic diversity of the varieties cultivated might actually increase if landrace replacement is only partial. We tested this hypothesis using microsatellite (SSR) markers to compare genetic diversity in the rice landraces and modern varieties of Kaski. We present the results of a follow-up to Joshi and Witcombe’s (2003) survey of farmers in Kaski to monitor changes in landrace cultivation after the introduction of M-3, M-9 and Lumle 2. 2. Materials and methods 2.1. Cultivation surveys Surveys were carried out in villages of the Kaski district (Fig. 1) in October 2003 and October 2004. Three of the villages were classed as mid-hill (between 1400 and 1600 m): Damdame, Kande and Marangche. Five were classed as high-hill (above 1600 m): Dhampus, Jhinje, Khanigaun, Tolka and Landruk. Three villages were classed as mountain (above 1600 m and with distinct physiographic features—chilling wind from the Himalayas and cold irrigation water): Chhomrong, Ghandruk-Athbhaihar and Chane. Individual farmers were interviewed and a knowledgeable person from each village accompanied the surveyors on a visit to the field for direct observations. Members of each farmers’ household were included in group discussions. An initial survey was made in five villages in 1996 (Sthapit et al., 1996) and this, along with surveys made in 1997, 1998 and 1999 (Joshi et al., 2001) were used to form the baseline data for 2003 and 2004 surveys. 2.2. Plant material Machhapuchhre-3 (M-3) and Machhapuchhre-9 (M-9) were derived from a cross between a local Kaski variety, Chhomrong Dhan with the Japanese variety Fuji 102 (japonica) that was made by the Division of Agricultural Botany, Nepal Agricultural Research Council for the Nepalese rice research programme. Lumle 2 was derived from a cross between Chhomrong Dhan and IR36 (indica) made at the Lumle Agricultural Research Centre. Henceforth we refer to M-3, M-9 and Lumle 2 as COB varieties (Table 1). Twelve rice landraces were selected for the analysis from those reported as changed or unchanged in frequency by Joshi and Witcombe (2003): six were randomly selected from the 10 that remained unchanged between 1996 and 1999 and six were randomly selected from the 8 that had shown a significant reduction in area. In subsequent surveys in 2003 and 2004 one landrace previously defined as unchanged had declined resulting in seven landraces in the declined category and five in the unchanged category. Seed samples of the landraces were collected from farmers in the villages of Dhikur Pokhari and Lumle in the
Fig. 1. Study sites in Kaski district (near Pokhara ^), Nepal: survey villages (*) and locations where rice landraces were collected (&).
Kaski district at elevations between 1280 and 1600 m in 2002 (Table 2). The 12 landraces, three COB varieties grown in Kaski (M-3, M-9 and Lumle 2) and six other modern varieties (checks) grown in other regions or countries (Fuji 102, IR36, IR64, Kalinga III, Radha 32 and Vandana) were sown in John Innes No. 2 compost in P84 trays (PlantPak, Maldon, Essex, UK) with partitions of 35 mm 35 mm 55 m deep. Seedlings were grown in a greenhouse at Pen-y-Ffridd field station, Bangor University, UK, under daylight supplemented by 150 Mol m2 s 1 PAR (minimum temperature 25 8C). Young leaf tissue from 2- to 3-week-old healthy seedlings was used for genomic DNA extraction. Leaves (100 mg) from 10 individual seedlings (pooled in equal quantities) of each accession were used for bulk DNA extractions with Qiagen DNeasy plant kits (Qiagen, Crawley, West Sussex, UK). DNA concentration was estimated using 0.8% agarose mini-gels run in 1 TBE buffer (0.09 M Tris–borate and 0.5 M EDTA) at 80 V for 90 min, with ethidium bromide staining. 2.3. Microsatellite analysis Forty-nine pairs of SSR primers (www.gramene.com) at loci dispersed throughout the rice genome were tested. Full details of
Table 1 Rice varieties developed with participatory client-oriented breeding (COB). Variety
Release date
Pedigree
Agronomic traits
Machhapuchhre-3 (M-3)
1996
LR88001-8C-OL Fuji 102/Chhomrong Dhan
Machhapuchhre-9 (M-9) Lumle-2 (Lumle 2)
Not yet released Not yet released
LR88001-18C-OL Fuji 102/Chhomrong Dhan IR36/Chhomrong Dhan
White grain, good cold tolerance, disease resistance (leaf and neck blast, bacterial sheath brown rot disease), maturity 174 d, high grain and straw yield, superior taste to Chhomrong; shattering problem As M-3 but moderate cold tolerance White grain, superior grain quality to M-3 and M-9, less shattering than M-3 and easier to thresh, good cold tolerance
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Table 2 Agronomic characteristics of 12 rice landraces (including landrace selection Chhomrong Dhan) cultivated in the Kaski district, Nepal and used for SSR analysis. Alternative names are given in brackets. Variety
Origin
Agronomic and other traits
Colour traits of grain
Maisare (Rato Takmare)
Local variety
Red grain
Kalopatle Raksali (Raksali, Galaiya) Kathe
Local variety Local variety from Lumle village Local variety (very old)
Silange
Local variety, a japonica type
Chhomrong Dhan
Marsi Juwari Rakse (Rause, Bhalu) Anga
Local released variety Pure line selection (1991) from Ghandruk Local, a japonica type Local variety Local variety Local variety Local variety
Suited to marginal conditions with limited water and nutrients, early maturing. Cold tolerant, low shattering, early maturing. Good yield potential for landrace, early maturing, poor taste. Moderately tolerant to cold, late maturing, low shattering, not responsive to fertilizers, good eating quality. Late maturing, cold tolerant, adapted to marginal conditions, coarse grain. Cold tolerant at reproductive stage, resistant to blast and sheath brown rot, tolerant to lodging.
White grain Dull white grain Dull white grain Red pericarp with long awns
Tarkaya (Tarkange) Panhele
Local variety Local variety
Late maturing, short round grain. Late maturing, moderately tolerant to cold. Short round grain. Suited to marginal conditions and limited water, shattering, low yield, good fodder, poor taste, medicinal value. Adapted to low fertility conditions, early maturing. High water-demanding, late maturing, good straw yield, low yield potential, susceptible to insect and disease, short round grain, aroma.
the PCR amplification protocol and fragment analysis on the CEQ 8000 (Beckman Coulter) were given in Steele et al. (2004).
White grain, dark glumes Dull white grain, yellow glumes White grain Dull white grain Red grain, speckled brown glumes
White grain, dark glumes, red apiculus
3. Results 3.1. Cultivation surveys
2.4. Data analysis Alleles detected in each variety were recorded in base pairs in a data matrix in MS Excel. The SimQual application of NTSYS-pc (Exeter Software) was used to find the Simple Matching Coefficient (SMC) matrix. The SMC matrix was used for cluster analysis (UPGMA clustering) and principal component analysis. The values in the SMC matrix for 5 landraces that did not decline from 1996 to 2004, i.e., non-declined landraces (ND LR), 7 landraces that declined from 1996 to 2004, i.e., declined landraces (D LR) and 3 COB varieties were used to develop a weighted diversity model (Witcombe, 1999) to determine the effect of different proportions of COB and landrace classes on total genetic diversity. The assumptions for modelling were: that within cultivar diversity was zero; that equal proportions of the varieties were grown within the COB class and each of the landrace classes; that the COB class varied from 0% to 100% of the area and that as the COB class increased the rate of decline for the D LR class was greater than the rate of decline for the ND LR class. The MS Excel Microsatellite Toolkit (http://www.animalgenomics.ucd.ie/sdepark/ms-toolkit/) was used to find Polymorphism Information Content, Nei’s unbiased gene diversity (Nei, 1987) and observed heterozygosity for each group of varieties.
The total area under cultivation with cold tolerant rice was 126 ha in the surveyed villages of Kaski. There were 754 recorded rice-growing households in 1996, of these 100 responded in 2003 (13%) and 131 in 2004 (17%). The average holding size per respondent household was 0.31 ha. The proportion of total area under modern COB varieties increased from zero (pre-1996) to approximately 60% by 2003 and 2004 (Fig. 2) when 49% of farmers only cultivated modern varieties. M-3 was the only COB variety adopted in the villages above 1600 m. In the lower altitude villages the COB varieties M-9 and Lumle 2 were more commonly taken up than M-3. The area under Lumle 2 increased at the expense of M-3 from 2003 to 2004. Twenty-five different landrace or local varieties were cultivated by at least one household in the survey since 1996. Fourteen of them were dropped by at least one farmer in favour of a COB variety between 1996 and 2004 (Table 3) and seven of these declined landraces were represented in the SSR study. Only one landrace was dropped by any farmer in any 1 year. Different varieties were dropped in different villages, for example Silange was only dropped in Chane village and Sinjali was dropped by all seven farmers surveyed in Damdame village. Chhomrong Dhan, a modern selection from a landrace, was the most frequently
Fig. 2. Proportion of area under cultivation of three modern COB varieties and rice landraces in 2003 and 2004, from results of farmer surveys in six high altitude villages of Kaski.
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Table 3 Rice landraces and local varieties recorded in surveys of farming households in Kaski since 1996. Alternative names are given in brackets. Variety
Change in number of households since 1996
Number of farmers who stated they had dropped it in 2004
Village(s) where dropped
Chhomrong Dhan
Declined 1996–1999
19
Kalo Dhana Lamjungea Sinjalia Kathe Silange Kalopatle Maisare (Rato Takmare) Maisare (Seto Takmare)a Tairingea Darmalia Reksali (Raksali; Galaiya) Seto Darmali (Darmili)a Rakse (Rause or Bhalu) Anga Galyamia Gharkholea Guntaa Juwari Khumal 4a Khumal 7a Marsi Pakhe Dhana Panhele Tarkaya (Tarkange)
Declined 1999–2004 Declined 1999–2004 Declined 1996–1999 Declined 1996–1999 Declined 1999–2004 Declined 1996–1999 Declined 1996–1999 Declined 1996–1999 Declined 1999–2004 Declined 1996–1999 Declined 1996–1999 Declined 1996–1999 Declined 1996–1999 No change by 2004 No change by 2004 No change by 2004 No change by 2004 No change by 2004 No change by 2004 No change by 2004 No change by 2004 No change by 2004 No change by 2004 No change by 2004
12 12 11 10 10 7 4 3 2 1 1 1
Chane, Chhomrong, Ghandruk-Athbhaithar, Jhinje, Khanigaun, Landruk, Tolka Chane, Jhinje Jhinje Chane, Khanigaun Marangche Damdame, Marangche Kande, Marangche Dhampus, Khanigaun Chane, Ghandruk-Athbhaithar Jhinje Chane Marangche Chane
a
Varieties not used for SSR analysis.
dropped variety. Chhomrong Dhan was a parent of all three COB varieties, so alleles from this variety remained in cultivation where COB varieties were adopted. 3.2. COB adoption in Marangche village In Marangche a higher proportion of households (75%, i.e. 25 from 33) were interviewed than in all other villages. In this village, the overall adoption of modern COB varieties by area was 33% in 2003 and 37% 2004. By 2004, Lumle 2 occupied a significant proportion (20%) of the whole of the village rice area even though there had been absolutely no outside intervention since its introduction. Lumle 2 area increased mainly at the expense of the previously released COB variety M-3. Analysis of individual households showed large differences in displacement levels. In 2003, 50% of the farmers had adopted the COB varieties on all of their rice land while 6% of farmers grew only landraces. Proportionally, there were more ‘100% COB adopters’ reported in Marangche than in any of the other surveyed villages. 3.3. Genetic diversity In total, 322 alleles were detected over 49 loci. As expected, the six genetically diverse check varieties showed high levels of polymorphism, having 170 different alleles of which 68 were not present in the landraces. The three COB varieties had 84 different alleles, of which 12 were not present in any of the landraces. A total of 239 different alleles were detected in the landraces and the mean PIC values for landraces and checks were similar at around 0.5 (Table 4). For the 12 landraces there was a mean of 4.9 1.9 alleles per locus and in the sub-set of five which had declined in popularity there was a mean of 3.8 1.6 alleles per locus (Table 5). The mean values for all three diversity measures (PIC, 1-SMC and Nei’s gene diversity) followed the same pattern: diversity of all 12 LR > diversity of 7 D LR > diversity of 5 ND LR > diversity of 3 COB (Tables 4 and 5).
Principal components analysis (PCA) with all varieties gave two separate clusters, one for the checks and one for all Kaski varieties (not shown). PCA performed without non-parent checks revealed three groups (Fig. 3). PCA axis 2 separated the three COB varieties and their parents from the other landraces. The landraces formed two clusters which had different members according to PCA axes 1 and 3. There was no relationship between clusters and landraces that declined. We modelled weighted genetic diversity of relative proportions of landrace classes and COB varieties. We assumed that the five non-declined landraces were most likely to persist as COB varieties become more popular, and so would decline less rapidly than the declined landraces class. Using these assumptions the model showed that the partial replacement of landraces led to an increase in genetic diversity up to adoption levels of COB of 65% (Fig. 4). Diversity progressively decreased only when levels of adoption exceeded 65%. 4. Discussion Joshi and Bauer (2007) expressed concern that the number of landraces grown by farmers in Nepal is declining, and although they conceded that landraces are not all of equal value to farming communities they called for public investment for the maintenance of unique ones. We have shown that although some landraces have been dropped completely the ones that are maintained contain a wide range of alleles. Therefore, we question the value of investment to preserve all of them. We argue that the overall benefit to the community from the uptake of COB varieties outweighs the loss of some landraces, and this is clearer when looking at the region as a whole. Our results are consistent with the overall findings of the genetic diversity studies reviewed by Fu (2006), i.e. that crop genetic diversity generally has not been significantly reduced by modern plant breeding, but that it has begun to decline. Although the COB varieties were adopted by more than 90% of farmers in Kaski, some on 100% of their land area, farmers across the district
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Table 4 SSR marker loci with chromosome location and polymorphism information content (PIC) values for all landraces (LR), only landraces that declined (D LR), only landraces that did not decline (ND LR), three modern varieties developed using client oriented breeding (COB) and modern check varieties (Checks). Locus
RM1 RM212 RM237 RM243 RM5 OSR14 RM110 RM211 RM213 RM221 RM233 RM262 RM279 RM29 RM318 RM48 RM6 RM16 RM22 RM347 RM55 RM241 RM349 RM13 RM153 RM163 RM188 RM26 RM413 RM440 RM593 RM3 RM528 Waxy RM11 RM2 RM234 RM248 RM351 RM51 RM223 RM350 RM105 RM242 RM342 RM239 RM258 RM206 RM229 Mean for group
Chromosome
PIC
1 1 1 1 1 2 2 2 2 2 2 2 2 2 2 2 2 3 3 3 3 4 4 5 5 5 5 5 5 5 5 6 6 6 7 7 7 7 7 7 8 8 9 9 9 10 10 11 11
All LR (n = 12)
D LR (n = 7)
ND LR (n = 5)
COB (n = 3)
Checks (n = 6)
0.60 0.21 0.65 0.85 0.64 0.61 0.79 0 0.39 0.59 0.54 0.46 0.57 0.58 0.28 0.72 0.28 0.30 0.66 0.62 0.57 0.81 0.63 0.74 0.61 0.81 0 0.54 0.62 0.64 0.60 0.80 0.60 0.64 0.66 0.60 0.58 0.45 0.28 0.57 0.61 0.44 0.64 0.15 0.73 0.29 0.39 0.84 0.81 0.55
0.41 0.12 0.37 0.86 0.55 0.60 0.79 0 0.41 0.58 0.46 0.35 0.67 0.52 0.24 0.55 0.24 0.28 0.57 0.62 0.50 0.73 0.52 0.52 0.64 0.73 0 0.53 0.41 0.59 0.41 0.78 0.52 0.57 0.62 0.50 0.33 0.57 0.21 0.58 0.67 0.32 0.64 0.24 0.72 0.45 0.41 0.82 0.70 0.50
0.57 0.27 0.50 0.67 0.49 0.54 0.73 0 0.27 0.50 0.50 0.54 0.27 0.56 0.31 0.64 0.27 0.33 0.65 0.54 0.56 0.60 0.67 0.67 0.47 0.61 0 0.55 0.50 0.55 0.50 0.50 0.64 0.50 0.45 0.45 0.50 0.16 0.33 0.45 0.27 0.55 0.50 0 0.38 0 0.27 0.70 0.69 0.45
0 0.35 0 0.35 0.35 0.35 0.45 0.35 0.35 0 0 0 0.35 0 0 0.35 0.35 0.35 0.35 0.67 0.35 0.35 0.59 0 0 0 0 0 0.35 0 0.35 0 0.35 0.35 0 0.35 0.35 0.35 0 0 0 0.45 0.35 0.35 0.54 0 0.35 0 0.35 0.22
0.24 0.76 0.60 0.50 0.62 0.50 0.24 0.27 0.60 0.45 0.56 0.73 0.54 0.24 0.55 0.45 0.62 0.62 0.36 0.53 0.42 0.55 0.54 0.45 0.54 0.54 0.35 0.46 0.62 0.54 0.45 0.27 0.24 0.69 0.66 0.36 0.69 0.73 0.35 0.48 0.64 0.45 0.67 0.66 0.67 0.45 0.74 0.54 0.60 0.52
continued to grow at least 11 diverse landraces on an average of 40% of the high-altitude land. On a global basis, Jarvis et al. (2008) found that considerable crop diversity was maintained on-farm in the form of traditional crop varieties. Landraces and local varieties are important for cultural reasons. For example, Chhomrong Dhan was most
frequently dropped by the farmers in our survey in favour of M3 (which shares 33% of its alleles with Chhomrong Dhan), yet it continued to be grown by a small number of farmers in the local Gurung community. For them it is the preferred rice for preparation of the dish Madeko Bhat used during funerals and other ritual and social ceremonies.
Table 5 Genetic diversity estimates from 49 SSR loci for all landraces, only landraces that declined (D LR), only landraces that did not change (ND LR), three modern varieties developed using client oriented breeding (COB) and modern check varieties (Checks).
All landraces D LR ND LR COB Checks a
Sample size
Genetic diversity (1-SMCa)
Nei’s unbiased gene diversity
Observed heterozygosities
Mean no. of alleles per locus
12 7 5 3 6
0.58 0.53 0.48 0.28 0.56
0.62 0.03 0.58 0.03 0.56 0.03 0.33 0.04 0.62 0.02
0.20 0.02 0.20 0.02 0.20 0.03 0.09 0.02 0.12 0.02
4.9 1.9 3.8 1.6 3.1 1.1 1.7 0.7 3.5 0.9
Simple Matching Coefficient.
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Fig. 4. Predicted changes in weighted genetic diversity as the proportion of COB varieties adopted increases relative to landraces. The arrow indicates the proportion of COB adoption above which diversity falls below the baseline level of no COB adoption. The model used values for 1-SMC values obtained for 12 landraces (seven D LR and five ND LR) and three COB varieties. We have assumed that the COB class varied from 0% to 100% of the area and that as the COB class increased the rate of decline for the D LR class was greater than the rate of decline for the ND LR class.
Fig. 3. PCA of Simple Matching Coefficients for the Kaski high altitude rice shows three groups containing: modern COB varieties (^), declined landraces (*), not declined landraces (*) and modern parent of COB, not grown in Kaski (^); (A) axis 1 axis 2; (B) axis 2 axis 3.
The COB varieties may also have contributed to additional allelic richness across the region as a whole (Fig. 4). We estimated genetic diversity would be greatest at adoption levels of COB at around 40% and that it would reduce below baseline levels only when adoption levels exceed 65%. The model assumes that varieties are grown in equal proportions in all villages, and clearly this assumption is an over-simplification. However, the model highlights that introducing modern varieties does not necessarily lead to an immediate decline in genetic diversity. This result supports the hypothesis based on Joshi and Witcombe (2003) that adoption of modern varieties bred from a cross between a local variety and an exotic variety can lead to an increase in the total allelic diversity if landrace replacement is only partial. There is likely to be a G E effect on the performance of individual modern varieties that influences their uptake by farmers. At altitudes below 1500 m M-9 may have slightly greater yields than M-3 while, at higher altitudes, the reverse applies because of the superior tolerance to chilling of M-3. Lumle 2, which was the last of the three COB varieties to reach farmers, is starting to become more popular than M-9 and local varieties at lower altitudes due to its superior grain quality and its lower panicle
shattering in the field at maturity. It also has better cold tolerance than M-9. We found greater levels of genetic diversity in landraces from Kaski than from the Jumla district of Nepal in a similar survey (Bajracharya et al., 2006). In Jumla, the only rice varieties that can be grown are those adapted to very high-altitude. The need for tolerance to extreme chilling stress creates a genetic bottleneck on the first introduction of any rice variety. In Kaski the lower altitudes remove the genetic bottleneck and allow an exchange of varieties between farmers that grow rice at a range of altitudes. The majority of cold tolerant rice varieties are japonicas. Several seedling cold tolerance QTLs have been mapped to chromosome 12 (Andaya and Tai, 2006 and references therein) however, none of the markers we tested were located on chromosome 12. Further work is necessary to test this landrace collection with more markers and it would be interesting to test for linkage disequilibrium around known cold tolerance QTL. 4.1. Conclusion The success of the COB programme conduced in Kaski for high altitude rice resulted in the rapid uptake of modern varieties whose cultivation improved the livelihoods of farming households. There was a corresponding reduction in the area and number of households growing some landraces but overall genetic diversity increased because the new varieties were genetically contrasting to the landraces. The new varieties had some alleles that were not present in any of the landraces and some alleles that might be lost in the displaced landraces. This emphasises the value of choosing locally adapted varieties as parents for breeding; it ensures that landrace genes are conserved and increases the likelihood that the breeding programme will succeed. Acknowledgements We thank Indra Poudel, Khem Choin, Bharat Bhandari, Jwala Bajracharya, Gwen Edwards and Julian Bridges for their valuable assistance in this work. The survey work was funded by Bioversity International and LI-BIRD. This document is an output from projects funded by the UK Department of International Development (DFID) and administered by CAZS-NR for the benefit of
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developing countries. The views expressed are not necessarily those of DFID. References Andaya, V.C., Tai, T.H., 2006. Fine mapping of the qCTS12 locus, a major QTL for seedling cold tolerance in rice. Theor. Appl. Genet. 113, 467–475. Bajracharya, J., Steele, K.A., Jarvis, D.I., Sthapit, B.R., Witcombe, J.R., 2006. Rice landrace diversity in Nepal: variability of agro-morphological traits and SSR markers in landraces from a high-altitude site. Field Crops Res. 95, 327– 335. Bardsley, D., Thomas, I., 2005. In situ agrobiodiversity conservation for regional development in Nepal. GeoJournal 62, 1572–9893. Brush, S.B., 2000. The issues of in situ conservation of crop genetic resources. In: Brush, S.B. (Ed.), Genes in the Field: On-farm Conservation of Crop Diversity. IPGRI, IDRC and Lewsis Pub, Washington, DC, pp. 3–26. Fu, Y.-B., 2006. Impact of plant breeding on genetic diversity of agricultural crops: searching for molecular evidence. Plant Genet. Resour. 4, 71–78. Jarvis, D.I., Brown, A.H.D., Cuong, P.H., Collado-Panduro, L., Latournerie-Moreno, L., Gyawali, S., Tanto, T., Sawadogo, M., Mar, I., Sadiki, M., Hue, N.T-N., Arias-Reyes, L., Balma, D., Bajracharya, J., Castillo, F., Rijal, D., Belqadi, L., Rana, R., Saidi, S., Ouedraogo, J., Zangre, R., Rhrib, K., Chavez, J.L., Schoen, D., Sthapit, B., De Santis, P., Fadda, C., Hodgkin, T., 2008. A global perspective of the richness and evenness of traditional crop-variety diversity maintained by farming communities. Proc. Natl. Acad. Sci. U.S.A. 105, 5326–5331. Joshi, G.R., Bauer, S., 2007. Cultivation and the loss of rice landraces in the Terai region of Nepal. Plant Genet. Resour. 5, 1–6.
Joshi, K.D., Sthapit, B.R., Gurung, R.B., Gurung, M.B., Witcombe, J.R., 1997. Machhapuchhre 3 (MP-3), the first rice variety developed through a participatory plant breeding approach released for mid to high altitudes of Nepal. IRRN 22 (2), 12. Joshi, K.D., Sthapit, B.R., Witcombe, J.R., 2001. How narrowly adapted are the products of decentralised breeding? The spread of rice varieties from a participatory plant breeding programme in Nepal. Euphytica 122, 589–597. Joshi, K.D., Witcombe, J.R., 2003. The impact of participatory plant breeding (PPB) on landrace diversity: a case study for high-altitude rice in Nepal. Euphytica 134, 117–125. Morris, M.L., Bellon, M.R., 2004. Participatory plant breeding research: opportunities and challenges for the international crop improvement system. Euphytica 136, 21–35. Nei, M., 1987. Molecular Evolutionary Genetics. Columbia University Press, New York, USA. Sthapit, B.R., 1992. Chilling injury of rice crop in Nepal: a review. J. Inst. Agric. Anim. Sci. 13, 1–32. Sthapit, B.R., Joshi, K.D., Witcombe, J.R., 1996. Farmer participatory cultivar improvement: a case of high altitude rice from Nepal. Exp. Agric. 32, 479–496. Steele, K.A., Edwards, G., Zhu, J., Witcombe, J.R., 2004. Marker-evaluated selection in rice: shifts in allele frequency among bulks selected in contrasting agricultural environments identify genomic regions of importance to rice adaptation and breeding. Theor. Appl. Genet. 109, 1247–1260. Witcombe, J.R., 1999. Does plant breeding lead to a loss of genetic diversity? In: Wood, D., Lenne, J.M. (Eds.), Agrobiodiversity Characterization, Utilization and Management. CAB International, Wallingford, UK, pp. 245–272. Witcombe, J.R., Joshi, K.D., Gyawali, S., Musa, A.M., Johansen, C., Virk, D.S., Sthapit, B.R., 2005. Participatory plant breeding is better described as highly clientoriented plant breeding. I. Four indicators of client-orientation in plant breeding. Exp. Agric. 41, 299–319.
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Has the introduction of modern rice varieties changed rice genetic diversity in a high-altitude region of Nepal? K.A. Steele a,*, S. Gyawali b, K.D. Joshi a, P. Shrestha b, B.R. Sthapit c, J.R. Witcombe a a
CAZS Natural Resources, Bangor University, Gwynedd LL57 2UW, UK LI-BIRD, PO Box 324, Giarapatan, Pokhara, Nepal c Bioversity International, Office for South Asia, National Agricultural Science Centre, DPS Marg, Pusa Campus, New Delhi 110012, India b
A R T I C L E I N F O
A B S T R A C T
Article history: Received 12 December 2008 Received in revised form 1 April 2009 Accepted 4 April 2009
We surveyed the uptake of three modern rice varieties by farmers in high-altitude villages in the Kaski district of Nepal and found that their uptake had displaced some traditional landraces in the district. The three varieties, Machhapuchhre-3 (M-3), Machhapuchhre-9 (M-9) and Lumle 2, were developed using client-oriented, participatory plant breeding methods and the first was introduced to farmers in 1996. By 2004 up to 60% of the land area was used to grow these modern varieties. Molecular markers (SSR) were used to assay levels of genetic diversity to test if adoption of modern varieties in the place of landraces had changed genetic diversity. The modern varieties were found to contain diverse alleles with a high proportion from the local parent variety, Chhomrong Dhan. We found a high level of allelic richness in the landraces, and although seven had been dropped in favour of the modern varieties, other diverse landraces were still being cultivated by farmers in the study villages on up to 40% of the rice area. Genetic diversity may be maintained even when landraces are displaced by modern varieties. Using a model we found that the partial replacement of landraces increased genetic diversity if the modern varieties were adopted on up to 65% of the area. Only above these levels did overall diversity decline. ß 2009 Elsevier B.V. All rights reserved.
Keywords: Client oriented breeding (COB) Participatory plant breeding (PPB) Rice Genetic diversity Genetic erosion Traditional variety Landrace Nepal
1. Introduction The adoption of modern varieties by farmers is a key development factor in Nepal that is starting to change cultivation practices and may promote genetic erosion (Bardsley and Thomas, 2005). In areas with harsh environments to which landraces are adapted genetic erosion could lead to the loss of important adaptive traits. An example is cold-tolerance in high altitude (1300–2200 m) rice landraces. The high-altitude rice area in Nepal is about 2% of the total 1.5 m ha rice growing area of Nepal, and until the mid-nineties very few modern varieties had been bred for these domains. Here cold injury due to cool air temperature is a major constraint to improving rice productivity (Sthapit, 1992) and plants are exposed to cold irrigation water (16–22 8C) during the reproductive phase. Three modern varieties for high altitudes of Nepal were developed in Kaski district through the successful use of clientoriented, participatory methods. Participatory plant breeding describes the activity (Morris and Bellon, 2004), whereas client oriented breeding (COB) describes its purpose (Witcombe et al.,
* Corresponding author. Tel.: +44 1248 388655; fax: +44 1248 370594. E-mail address: [email protected] (K.A. Steele). 0378-4290/$ – see front matter ß 2009 Elsevier B.V. All rights reserved. doi:10.1016/j.fcr.2009.04.002
2005). This breeding programme led to the release of Machhapuchhre-3 (M-3) in 1997 (Sthapit et al., 1996; Joshi et al., 1997). Subsequently a sister line Machhapuchhre-9 (M-9) was identified as being highly desirable (Joshi et al., 2001). In 2001, a third modern variety, Lumle 2, from a different cross was identified as suitable by farmers in high altitude villages in Kaski. Although not yet formally released, both M-9 and Lumle 2 have been adopted by farmers and are spreading through farmer-to-farmer seed networks. The effects of introducing modern varieties on changes in the cultivation of rice landraces between 1996 and 1999 in highaltitude regions of Kaski were recorded by Joshi et al. (2001) and Joshi and Witcombe (2003). Landrace cultivation decreased as a proportion of the total area under cultivation and by fewer households growing a given landrace, however few landraces were dropped completely. The adoption of modern varieties caused a greater decline in the more commonly grown landraces while the less common ones were not replaced so rapidly because farmers kept them for their specific adaptations and uses. Participatory breeding methods are intended to produce better quality and higher yielding varieties that are adapted to local cultivation, however there are widespread concerns that the uptake of modern varieties may have negative impacts on genetic diversity (Brush, 2000). Witcombe (1999) has argued that such
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breeding methods do not necessarily lead to a loss of genetic diversity, and that use of participatory methods may even widen crop genetic diversity, particularly where existing biodiversity in farmers’ fields is low. We hypothesised that by adopting modern varieties bred from a cross between a local variety and an exotic variety, the total allelic diversity of the varieties cultivated might actually increase if landrace replacement is only partial. We tested this hypothesis using microsatellite (SSR) markers to compare genetic diversity in the rice landraces and modern varieties of Kaski. We present the results of a follow-up to Joshi and Witcombe’s (2003) survey of farmers in Kaski to monitor changes in landrace cultivation after the introduction of M-3, M-9 and Lumle 2. 2. Materials and methods 2.1. Cultivation surveys Surveys were carried out in villages of the Kaski district (Fig. 1) in October 2003 and October 2004. Three of the villages were classed as mid-hill (between 1400 and 1600 m): Damdame, Kande and Marangche. Five were classed as high-hill (above 1600 m): Dhampus, Jhinje, Khanigaun, Tolka and Landruk. Three villages were classed as mountain (above 1600 m and with distinct physiographic features—chilling wind from the Himalayas and cold irrigation water): Chhomrong, Ghandruk-Athbhaihar and Chane. Individual farmers were interviewed and a knowledgeable person from each village accompanied the surveyors on a visit to the field for direct observations. Members of each farmers’ household were included in group discussions. An initial survey was made in five villages in 1996 (Sthapit et al., 1996) and this, along with surveys made in 1997, 1998 and 1999 (Joshi et al., 2001) were used to form the baseline data for 2003 and 2004 surveys. 2.2. Plant material Machhapuchhre-3 (M-3) and Machhapuchhre-9 (M-9) were derived from a cross between a local Kaski variety, Chhomrong Dhan with the Japanese variety Fuji 102 (japonica) that was made by the Division of Agricultural Botany, Nepal Agricultural Research Council for the Nepalese rice research programme. Lumle 2 was derived from a cross between Chhomrong Dhan and IR36 (indica) made at the Lumle Agricultural Research Centre. Henceforth we refer to M-3, M-9 and Lumle 2 as COB varieties (Table 1). Twelve rice landraces were selected for the analysis from those reported as changed or unchanged in frequency by Joshi and Witcombe (2003): six were randomly selected from the 10 that remained unchanged between 1996 and 1999 and six were randomly selected from the 8 that had shown a significant reduction in area. In subsequent surveys in 2003 and 2004 one landrace previously defined as unchanged had declined resulting in seven landraces in the declined category and five in the unchanged category. Seed samples of the landraces were collected from farmers in the villages of Dhikur Pokhari and Lumle in the
Fig. 1. Study sites in Kaski district (near Pokhara ^), Nepal: survey villages (*) and locations where rice landraces were collected (&).
Kaski district at elevations between 1280 and 1600 m in 2002 (Table 2). The 12 landraces, three COB varieties grown in Kaski (M-3, M-9 and Lumle 2) and six other modern varieties (checks) grown in other regions or countries (Fuji 102, IR36, IR64, Kalinga III, Radha 32 and Vandana) were sown in John Innes No. 2 compost in P84 trays (PlantPak, Maldon, Essex, UK) with partitions of 35 mm 35 mm 55 m deep. Seedlings were grown in a greenhouse at Pen-y-Ffridd field station, Bangor University, UK, under daylight supplemented by 150 Mol m2 s 1 PAR (minimum temperature 25 8C). Young leaf tissue from 2- to 3-week-old healthy seedlings was used for genomic DNA extraction. Leaves (100 mg) from 10 individual seedlings (pooled in equal quantities) of each accession were used for bulk DNA extractions with Qiagen DNeasy plant kits (Qiagen, Crawley, West Sussex, UK). DNA concentration was estimated using 0.8% agarose mini-gels run in 1 TBE buffer (0.09 M Tris–borate and 0.5 M EDTA) at 80 V for 90 min, with ethidium bromide staining. 2.3. Microsatellite analysis Forty-nine pairs of SSR primers (www.gramene.com) at loci dispersed throughout the rice genome were tested. Full details of
Table 1 Rice varieties developed with participatory client-oriented breeding (COB). Variety
Release date
Pedigree
Agronomic traits
Machhapuchhre-3 (M-3)
1996
LR88001-8C-OL Fuji 102/Chhomrong Dhan
Machhapuchhre-9 (M-9) Lumle-2 (Lumle 2)
Not yet released Not yet released
LR88001-18C-OL Fuji 102/Chhomrong Dhan IR36/Chhomrong Dhan
White grain, good cold tolerance, disease resistance (leaf and neck blast, bacterial sheath brown rot disease), maturity 174 d, high grain and straw yield, superior taste to Chhomrong; shattering problem As M-3 but moderate cold tolerance White grain, superior grain quality to M-3 and M-9, less shattering than M-3 and easier to thresh, good cold tolerance
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Table 2 Agronomic characteristics of 12 rice landraces (including landrace selection Chhomrong Dhan) cultivated in the Kaski district, Nepal and used for SSR analysis. Alternative names are given in brackets. Variety
Origin
Agronomic and other traits
Colour traits of grain
Maisare (Rato Takmare)
Local variety
Red grain
Kalopatle Raksali (Raksali, Galaiya) Kathe
Local variety Local variety from Lumle village Local variety (very old)
Silange
Local variety, a japonica type
Chhomrong Dhan
Marsi Juwari Rakse (Rause, Bhalu) Anga
Local released variety Pure line selection (1991) from Ghandruk Local, a japonica type Local variety Local variety Local variety Local variety
Suited to marginal conditions with limited water and nutrients, early maturing. Cold tolerant, low shattering, early maturing. Good yield potential for landrace, early maturing, poor taste. Moderately tolerant to cold, late maturing, low shattering, not responsive to fertilizers, good eating quality. Late maturing, cold tolerant, adapted to marginal conditions, coarse grain. Cold tolerant at reproductive stage, resistant to blast and sheath brown rot, tolerant to lodging.
White grain Dull white grain Dull white grain Red pericarp with long awns
Tarkaya (Tarkange) Panhele
Local variety Local variety
Late maturing, short round grain. Late maturing, moderately tolerant to cold. Short round grain. Suited to marginal conditions and limited water, shattering, low yield, good fodder, poor taste, medicinal value. Adapted to low fertility conditions, early maturing. High water-demanding, late maturing, good straw yield, low yield potential, susceptible to insect and disease, short round grain, aroma.
the PCR amplification protocol and fragment analysis on the CEQ 8000 (Beckman Coulter) were given in Steele et al. (2004).
White grain, dark glumes Dull white grain, yellow glumes White grain Dull white grain Red grain, speckled brown glumes
White grain, dark glumes, red apiculus
3. Results 3.1. Cultivation surveys
2.4. Data analysis Alleles detected in each variety were recorded in base pairs in a data matrix in MS Excel. The SimQual application of NTSYS-pc (Exeter Software) was used to find the Simple Matching Coefficient (SMC) matrix. The SMC matrix was used for cluster analysis (UPGMA clustering) and principal component analysis. The values in the SMC matrix for 5 landraces that did not decline from 1996 to 2004, i.e., non-declined landraces (ND LR), 7 landraces that declined from 1996 to 2004, i.e., declined landraces (D LR) and 3 COB varieties were used to develop a weighted diversity model (Witcombe, 1999) to determine the effect of different proportions of COB and landrace classes on total genetic diversity. The assumptions for modelling were: that within cultivar diversity was zero; that equal proportions of the varieties were grown within the COB class and each of the landrace classes; that the COB class varied from 0% to 100% of the area and that as the COB class increased the rate of decline for the D LR class was greater than the rate of decline for the ND LR class. The MS Excel Microsatellite Toolkit (http://www.animalgenomics.ucd.ie/sdepark/ms-toolkit/) was used to find Polymorphism Information Content, Nei’s unbiased gene diversity (Nei, 1987) and observed heterozygosity for each group of varieties.
The total area under cultivation with cold tolerant rice was 126 ha in the surveyed villages of Kaski. There were 754 recorded rice-growing households in 1996, of these 100 responded in 2003 (13%) and 131 in 2004 (17%). The average holding size per respondent household was 0.31 ha. The proportion of total area under modern COB varieties increased from zero (pre-1996) to approximately 60% by 2003 and 2004 (Fig. 2) when 49% of farmers only cultivated modern varieties. M-3 was the only COB variety adopted in the villages above 1600 m. In the lower altitude villages the COB varieties M-9 and Lumle 2 were more commonly taken up than M-3. The area under Lumle 2 increased at the expense of M-3 from 2003 to 2004. Twenty-five different landrace or local varieties were cultivated by at least one household in the survey since 1996. Fourteen of them were dropped by at least one farmer in favour of a COB variety between 1996 and 2004 (Table 3) and seven of these declined landraces were represented in the SSR study. Only one landrace was dropped by any farmer in any 1 year. Different varieties were dropped in different villages, for example Silange was only dropped in Chane village and Sinjali was dropped by all seven farmers surveyed in Damdame village. Chhomrong Dhan, a modern selection from a landrace, was the most frequently
Fig. 2. Proportion of area under cultivation of three modern COB varieties and rice landraces in 2003 and 2004, from results of farmer surveys in six high altitude villages of Kaski.
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Table 3 Rice landraces and local varieties recorded in surveys of farming households in Kaski since 1996. Alternative names are given in brackets. Variety
Change in number of households since 1996
Number of farmers who stated they had dropped it in 2004
Village(s) where dropped
Chhomrong Dhan
Declined 1996–1999
19
Kalo Dhana Lamjungea Sinjalia Kathe Silange Kalopatle Maisare (Rato Takmare) Maisare (Seto Takmare)a Tairingea Darmalia Reksali (Raksali; Galaiya) Seto Darmali (Darmili)a Rakse (Rause or Bhalu) Anga Galyamia Gharkholea Guntaa Juwari Khumal 4a Khumal 7a Marsi Pakhe Dhana Panhele Tarkaya (Tarkange)
Declined 1999–2004 Declined 1999–2004 Declined 1996–1999 Declined 1996–1999 Declined 1999–2004 Declined 1996–1999 Declined 1996–1999 Declined 1996–1999 Declined 1999–2004 Declined 1996–1999 Declined 1996–1999 Declined 1996–1999 Declined 1996–1999 No change by 2004 No change by 2004 No change by 2004 No change by 2004 No change by 2004 No change by 2004 No change by 2004 No change by 2004 No change by 2004 No change by 2004 No change by 2004
12 12 11 10 10 7 4 3 2 1 1 1
Chane, Chhomrong, Ghandruk-Athbhaithar, Jhinje, Khanigaun, Landruk, Tolka Chane, Jhinje Jhinje Chane, Khanigaun Marangche Damdame, Marangche Kande, Marangche Dhampus, Khanigaun Chane, Ghandruk-Athbhaithar Jhinje Chane Marangche Chane
a
Varieties not used for SSR analysis.
dropped variety. Chhomrong Dhan was a parent of all three COB varieties, so alleles from this variety remained in cultivation where COB varieties were adopted. 3.2. COB adoption in Marangche village In Marangche a higher proportion of households (75%, i.e. 25 from 33) were interviewed than in all other villages. In this village, the overall adoption of modern COB varieties by area was 33% in 2003 and 37% 2004. By 2004, Lumle 2 occupied a significant proportion (20%) of the whole of the village rice area even though there had been absolutely no outside intervention since its introduction. Lumle 2 area increased mainly at the expense of the previously released COB variety M-3. Analysis of individual households showed large differences in displacement levels. In 2003, 50% of the farmers had adopted the COB varieties on all of their rice land while 6% of farmers grew only landraces. Proportionally, there were more ‘100% COB adopters’ reported in Marangche than in any of the other surveyed villages. 3.3. Genetic diversity In total, 322 alleles were detected over 49 loci. As expected, the six genetically diverse check varieties showed high levels of polymorphism, having 170 different alleles of which 68 were not present in the landraces. The three COB varieties had 84 different alleles, of which 12 were not present in any of the landraces. A total of 239 different alleles were detected in the landraces and the mean PIC values for landraces and checks were similar at around 0.5 (Table 4). For the 12 landraces there was a mean of 4.9 1.9 alleles per locus and in the sub-set of five which had declined in popularity there was a mean of 3.8 1.6 alleles per locus (Table 5). The mean values for all three diversity measures (PIC, 1-SMC and Nei’s gene diversity) followed the same pattern: diversity of all 12 LR > diversity of 7 D LR > diversity of 5 ND LR > diversity of 3 COB (Tables 4 and 5).
Principal components analysis (PCA) with all varieties gave two separate clusters, one for the checks and one for all Kaski varieties (not shown). PCA performed without non-parent checks revealed three groups (Fig. 3). PCA axis 2 separated the three COB varieties and their parents from the other landraces. The landraces formed two clusters which had different members according to PCA axes 1 and 3. There was no relationship between clusters and landraces that declined. We modelled weighted genetic diversity of relative proportions of landrace classes and COB varieties. We assumed that the five non-declined landraces were most likely to persist as COB varieties become more popular, and so would decline less rapidly than the declined landraces class. Using these assumptions the model showed that the partial replacement of landraces led to an increase in genetic diversity up to adoption levels of COB of 65% (Fig. 4). Diversity progressively decreased only when levels of adoption exceeded 65%. 4. Discussion Joshi and Bauer (2007) expressed concern that the number of landraces grown by farmers in Nepal is declining, and although they conceded that landraces are not all of equal value to farming communities they called for public investment for the maintenance of unique ones. We have shown that although some landraces have been dropped completely the ones that are maintained contain a wide range of alleles. Therefore, we question the value of investment to preserve all of them. We argue that the overall benefit to the community from the uptake of COB varieties outweighs the loss of some landraces, and this is clearer when looking at the region as a whole. Our results are consistent with the overall findings of the genetic diversity studies reviewed by Fu (2006), i.e. that crop genetic diversity generally has not been significantly reduced by modern plant breeding, but that it has begun to decline. Although the COB varieties were adopted by more than 90% of farmers in Kaski, some on 100% of their land area, farmers across the district
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Table 4 SSR marker loci with chromosome location and polymorphism information content (PIC) values for all landraces (LR), only landraces that declined (D LR), only landraces that did not decline (ND LR), three modern varieties developed using client oriented breeding (COB) and modern check varieties (Checks). Locus
RM1 RM212 RM237 RM243 RM5 OSR14 RM110 RM211 RM213 RM221 RM233 RM262 RM279 RM29 RM318 RM48 RM6 RM16 RM22 RM347 RM55 RM241 RM349 RM13 RM153 RM163 RM188 RM26 RM413 RM440 RM593 RM3 RM528 Waxy RM11 RM2 RM234 RM248 RM351 RM51 RM223 RM350 RM105 RM242 RM342 RM239 RM258 RM206 RM229 Mean for group
Chromosome
PIC
1 1 1 1 1 2 2 2 2 2 2 2 2 2 2 2 2 3 3 3 3 4 4 5 5 5 5 5 5 5 5 6 6 6 7 7 7 7 7 7 8 8 9 9 9 10 10 11 11
All LR (n = 12)
D LR (n = 7)
ND LR (n = 5)
COB (n = 3)
Checks (n = 6)
0.60 0.21 0.65 0.85 0.64 0.61 0.79 0 0.39 0.59 0.54 0.46 0.57 0.58 0.28 0.72 0.28 0.30 0.66 0.62 0.57 0.81 0.63 0.74 0.61 0.81 0 0.54 0.62 0.64 0.60 0.80 0.60 0.64 0.66 0.60 0.58 0.45 0.28 0.57 0.61 0.44 0.64 0.15 0.73 0.29 0.39 0.84 0.81 0.55
0.41 0.12 0.37 0.86 0.55 0.60 0.79 0 0.41 0.58 0.46 0.35 0.67 0.52 0.24 0.55 0.24 0.28 0.57 0.62 0.50 0.73 0.52 0.52 0.64 0.73 0 0.53 0.41 0.59 0.41 0.78 0.52 0.57 0.62 0.50 0.33 0.57 0.21 0.58 0.67 0.32 0.64 0.24 0.72 0.45 0.41 0.82 0.70 0.50
0.57 0.27 0.50 0.67 0.49 0.54 0.73 0 0.27 0.50 0.50 0.54 0.27 0.56 0.31 0.64 0.27 0.33 0.65 0.54 0.56 0.60 0.67 0.67 0.47 0.61 0 0.55 0.50 0.55 0.50 0.50 0.64 0.50 0.45 0.45 0.50 0.16 0.33 0.45 0.27 0.55 0.50 0 0.38 0 0.27 0.70 0.69 0.45
0 0.35 0 0.35 0.35 0.35 0.45 0.35 0.35 0 0 0 0.35 0 0 0.35 0.35 0.35 0.35 0.67 0.35 0.35 0.59 0 0 0 0 0 0.35 0 0.35 0 0.35 0.35 0 0.35 0.35 0.35 0 0 0 0.45 0.35 0.35 0.54 0 0.35 0 0.35 0.22
0.24 0.76 0.60 0.50 0.62 0.50 0.24 0.27 0.60 0.45 0.56 0.73 0.54 0.24 0.55 0.45 0.62 0.62 0.36 0.53 0.42 0.55 0.54 0.45 0.54 0.54 0.35 0.46 0.62 0.54 0.45 0.27 0.24 0.69 0.66 0.36 0.69 0.73 0.35 0.48 0.64 0.45 0.67 0.66 0.67 0.45 0.74 0.54 0.60 0.52
continued to grow at least 11 diverse landraces on an average of 40% of the high-altitude land. On a global basis, Jarvis et al. (2008) found that considerable crop diversity was maintained on-farm in the form of traditional crop varieties. Landraces and local varieties are important for cultural reasons. For example, Chhomrong Dhan was most
frequently dropped by the farmers in our survey in favour of M3 (which shares 33% of its alleles with Chhomrong Dhan), yet it continued to be grown by a small number of farmers in the local Gurung community. For them it is the preferred rice for preparation of the dish Madeko Bhat used during funerals and other ritual and social ceremonies.
Table 5 Genetic diversity estimates from 49 SSR loci for all landraces, only landraces that declined (D LR), only landraces that did not change (ND LR), three modern varieties developed using client oriented breeding (COB) and modern check varieties (Checks).
All landraces D LR ND LR COB Checks a
Sample size
Genetic diversity (1-SMCa)
Nei’s unbiased gene diversity
Observed heterozygosities
Mean no. of alleles per locus
12 7 5 3 6
0.58 0.53 0.48 0.28 0.56
0.62 0.03 0.58 0.03 0.56 0.03 0.33 0.04 0.62 0.02
0.20 0.02 0.20 0.02 0.20 0.03 0.09 0.02 0.12 0.02
4.9 1.9 3.8 1.6 3.1 1.1 1.7 0.7 3.5 0.9
Simple Matching Coefficient.
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Fig. 4. Predicted changes in weighted genetic diversity as the proportion of COB varieties adopted increases relative to landraces. The arrow indicates the proportion of COB adoption above which diversity falls below the baseline level of no COB adoption. The model used values for 1-SMC values obtained for 12 landraces (seven D LR and five ND LR) and three COB varieties. We have assumed that the COB class varied from 0% to 100% of the area and that as the COB class increased the rate of decline for the D LR class was greater than the rate of decline for the ND LR class.
Fig. 3. PCA of Simple Matching Coefficients for the Kaski high altitude rice shows three groups containing: modern COB varieties (^), declined landraces (*), not declined landraces (*) and modern parent of COB, not grown in Kaski (^); (A) axis 1 axis 2; (B) axis 2 axis 3.
The COB varieties may also have contributed to additional allelic richness across the region as a whole (Fig. 4). We estimated genetic diversity would be greatest at adoption levels of COB at around 40% and that it would reduce below baseline levels only when adoption levels exceed 65%. The model assumes that varieties are grown in equal proportions in all villages, and clearly this assumption is an over-simplification. However, the model highlights that introducing modern varieties does not necessarily lead to an immediate decline in genetic diversity. This result supports the hypothesis based on Joshi and Witcombe (2003) that adoption of modern varieties bred from a cross between a local variety and an exotic variety can lead to an increase in the total allelic diversity if landrace replacement is only partial. There is likely to be a G E effect on the performance of individual modern varieties that influences their uptake by farmers. At altitudes below 1500 m M-9 may have slightly greater yields than M-3 while, at higher altitudes, the reverse applies because of the superior tolerance to chilling of M-3. Lumle 2, which was the last of the three COB varieties to reach farmers, is starting to become more popular than M-9 and local varieties at lower altitudes due to its superior grain quality and its lower panicle
shattering in the field at maturity. It also has better cold tolerance than M-9. We found greater levels of genetic diversity in landraces from Kaski than from the Jumla district of Nepal in a similar survey (Bajracharya et al., 2006). In Jumla, the only rice varieties that can be grown are those adapted to very high-altitude. The need for tolerance to extreme chilling stress creates a genetic bottleneck on the first introduction of any rice variety. In Kaski the lower altitudes remove the genetic bottleneck and allow an exchange of varieties between farmers that grow rice at a range of altitudes. The majority of cold tolerant rice varieties are japonicas. Several seedling cold tolerance QTLs have been mapped to chromosome 12 (Andaya and Tai, 2006 and references therein) however, none of the markers we tested were located on chromosome 12. Further work is necessary to test this landrace collection with more markers and it would be interesting to test for linkage disequilibrium around known cold tolerance QTL. 4.1. Conclusion The success of the COB programme conduced in Kaski for high altitude rice resulted in the rapid uptake of modern varieties whose cultivation improved the livelihoods of farming households. There was a corresponding reduction in the area and number of households growing some landraces but overall genetic diversity increased because the new varieties were genetically contrasting to the landraces. The new varieties had some alleles that were not present in any of the landraces and some alleles that might be lost in the displaced landraces. This emphasises the value of choosing locally adapted varieties as parents for breeding; it ensures that landrace genes are conserved and increases the likelihood that the breeding programme will succeed. Acknowledgements We thank Indra Poudel, Khem Choin, Bharat Bhandari, Jwala Bajracharya, Gwen Edwards and Julian Bridges for their valuable assistance in this work. The survey work was funded by Bioversity International and LI-BIRD. This document is an output from projects funded by the UK Department of International Development (DFID) and administered by CAZS-NR for the benefit of
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