The effectiveness of evaluating wild species: searching for sources of resistance to bruchid beetles in the genus Vigna subgenus Ceratotropis

A species level germplasm collection representing 76% of known taxa in the genus Vigna subgenus Ceratotropis was evaluated for resistance to two species of bruchid beetles, Callosobruchus chinensis and C. maculatus. Seven taxa consisting of 29 accessions were found to be resistant to C. chinensis and 4 taxa consisting of 24 accessions were found to be resistant to C. maculatus. This compared with no resistant accessions being found in several hundred landrace accessions of mungbean, V. radiata var. radiata, in the same subgenus. Sometimes resistance was found in all accessions of a particular taxon, such as complete resistance to both C. chinensis and C. macualtus in V. umbellata. Other taxa showed intra taxon variation for resistance such as V. reflexo-pilosa andV. minima. The levels and patterns of resistance among taxa were diverse. The results suggest that various factors cause resistance to bruchid in the subgenus Ceratotropis. While the number of eggs laid on seeds generally reflected seed size, one small seeded cultivar of V. mungo var. mungo, black gram, had an unusually high number of eggs laid per seed. No correlation was found between seed size and levels of resistance. The species level germplasm collection, which reflects the core collection concept in trying to maximize genetic diversity in a limited number of accessions, has enabled a large number of potentially useful sources of resistance to bruchid beetles to be found efficiently. This revised version was published online in July 2006 with corrections to the Cover Date.     

Morphological and molecular characterization of melon accessions resistant to Fusarium wilts

Fusarium wilt incited by Fusarium oxysporum f. sp. melonis (F.o.m) is one of the most widespread and devastating melon diseases. While resistance to physiological races 0, 1, and 2 is relatively frequent in different botanical varieties, sources of resistance to race 1,2 are restricted to a few Far-Eastern accessions. In this work, the results of a screening for resistance to F.o.m. race 1,2 among 32 accessions are presented. Three Japanese accessions (‘Kogane Nashi Makuwa’, ‘C-211’, and ‘C-40’) showed the highest resistance levels, but useful levels of resistance were also detected in one Russian ‘C-160’ and two Spanish (‘C-300’ and ‘Mollerusa-7’) accessions. These resistant materials, together with other accessions previously described as resistant to F.o.m. races 0, 1, and/or 2 have been morphologically and molecularly characterized. Based on cluster analysis, these accessions have been grouped according to the botanical subspecies they belong to. Assessment of genetic diversity indicated that the resistant accessions to races 0, 1 and 2, are scattered along the established clusters. On the other hand, high levels of resistance to the race 1,2 could be found only among accessions belonging to Cucumis melo subsp. agrestis, nevertheless, a certain degree of resistance to this race could also be found within some accessions belonging to subsp. melo. As far as we know, this is the first report of resistance to F.o.m race 1,2 found out from the Far-Eastern melon material. Based on fruits characteristics, it appears that several inodurus and cantalupensis accessions could be exploited in breeding programs as resistance sources to F.o.m races 0, 1 and/or 2 for the improvement of these melon types. The accessions with the highest levels of resistance to the race 1,2 appeared to be very distant both molecularly and morphologically from the commercial types. Nevertheless ‘C-160’, ‘C-300’, and ‘Mollerusa-7’ classified as var. inodorus are morphologically very similar to the Spanish commercial types and might be used as resistant sources in breeding these melon types.     

Immunity to Clavibacter michiganensis subsp. sepedonicus: Screening of exotic Solanum species

Summary Accessions from exotic Solanum species, including diploid and tetraploid species, were screened for immunity to Clavibacter michiganensis subsp. sepedonicus, the causal agent of potato ring rot. The diploid species included S. infundibuliforme, S. lesteri, S. megistacrolobum, S. tuberosum Group Phureja, S. polyadenium, S. pinnatisectum, S. raphanifolium, S. sparsipilum, S. sanctae-rosae, S. tuberosum Group Stenotomum, S. toralapanum, and S. verrucosum. The tetraploid species included S. tuberosum Group Andigena, S. acaule, S. fendleri, S. hjertingii, S. oplocense, S. polytrichon, and S. stoloniferum. Apparent immunity was initially found in several diploid species, but was not present during subsequent retesting. Immunity was found in nine accessions of tetraploid S. acaule. These accessions maintained their immunity during testing over an eight-month period. S. acaule appears to be a good source of immunity for introgression studies.     

Sources of resistance to diseases of sugar beet in related <Emphasis Type="Italic">Beta</Emphasis> germplasm: I. Foliar diseases

Resistance to four foliar diseases of sugar beet (Beta vulgaris ssp. vulgaris), virus yellows caused by Beet mild yellowing virus (BMYV) and Beet yellows virus (BYV), powdery mildew (Erysiphe betae) and Cercospora leaf spot (Cercospora beticola), was assessed in up to 600 accessions of closely related wild and cultivated Beta species. Most accessions were from the Section Beta, a taxon containing types most closely related to, and sexually compatible with, sugar beet and therefore most valuable for use in crop improvement. Between 1–12% of accessions were highly resistant (resistance scores of 2 on an international standardised resistance scale of 1–9) to these diseases. These levels, however, underestimate the potential number of resistant sources available from this section as some accessions with intermediate mean resistance scores contained a significant proportion of highly resistant plants within segregating populations. Variation in resistance to all diseases except BYV was observed within the Section Beta. Much higher levels of resistance were observed, and more frequently, in more distantly related sections of the genus Beta. Accessions of the Section Corollinae were highly resistant to both viruses (>62% of accessions tested), but less so to Cercospora (15%) and they were very susceptible to powdery mildew. Section Procumbentes accessions were highly resistant to BMYV and Cercospora (100%) but less so to powdery mildew (50%) and BYV (20%). However, sexual incompatibility between these sections and sugar beet make utilisation of these sources impractical using conventional breeding methods.     

RAPD analysis of interspecific relationships in presumably apomictic Cotoneaster species

A few of the approximately 300 Cotoneaster species described are diploid but the majority appear to be polyploid. The occurrence of apomixis inpolyploid Cotoneaster species has been reported but never proven with genetic markers. We have used 76 polymorphic RAPD markers to investigate the breeding system and phenetic grouping of some critical taxa, including a total of 19 plant accessions representing 13 mostly European species in the series Cotoneaster. Three to four individual plants, raised from seed from the same original plant, were analyzed for each of three accessions to investigate the possible occurrence of apomictic seed set. Absolutely congruent RAPD profiles were encountered among seedlings from one accession, whereas we found one or two marker differences among seedlings from the other two accessions. Genetic similarities among the different accessions were analyzed with a UPGMA-derived dendrogram. The most deviant taxon was the Chinese C. albokermesinus. A group withC. soczavianus and C. tomentosus was rather isolated from the remainder, as was also C. kullensis. Among the remaining taxa, two well supported clusters were found: (1) C. antoninae and C. uralensis, and (2) C. integerrimus and C. raboutensis, whereas the other five species (C. canescens, C. niger, C. scandinavicus, C. juranus, C. cambricus) formed a poorly supported cluster with no clear substructuring. A principal coordinate analysis yielded results that were in good correspondence with the dendrogram. Again C. albokermesinus appears to be totally isolated from the other species. In addition, two well-defined and rather isolated groups were found: (1) C. tomentosus and C. soczavianus, and (2) C. antoninae and C. uralensis, with the remainder comprising a loosely defined group. This revised version was published online in July 2006 with corrections to the Cover Date.     

Genetic diversity and taxonomic relationships within the genus Lens as revealed by allozyme polymorphism

Summary A survey of allozyme polymorphism at 11 loci was carried out on 439 accessions from the genus Lens. This comprised 153 Lens culinaris subsp. orientalis, 35 L. odemensis, 117 L. ervoides, 32 L. nigricans, 2 of a differentiated cytotype of L. nigricans and 100 landrace accessions of the cultivated lentil (L. culinaris subsp. culinaris), from 10 different countries. The aim of the survey was to determine intra-specific genetic diversity and species relationships, based on phylogenetic and phenetic analyses, particularly regarding the position of L. odemensis and the differentiated cytotype of L. nigricans. Diversity was described by three statistics. The level of diversity in the cultivated taxon was lower than in any of the wild species according to two of these statistics, the percentage of polymorphic loci and mean number of alleles per locus. For the third measure (Nei's mean genetic diversity) it was only greater than L. ervoides. Genetic diversity statistics of the wild species indicated differences in the nature of between-population genetic diversity within the different taxa. Phylogenetic analysis suggests that L. odemensis and L. ervoides evolved from a common ancestor, and L. culinaris subsp. orientalis subsequently evolved from L. odemensis. Phenetic analysis, however, places L. odemensis closer to L. culinaris subsp. orientalis than to L. ervoides. Nei's mean genetic distance of L. odemensis from both L. culinaris subsp. culinaris (0.204) and L. culinaris subsp. orientalis (0.110) was greater than the distance between them (0.062). This evidence is not conclusive in determining whether L. odemensis should retain its specific status. Further crossability studies should be carried out on a range of genotypes to assess the potential for gene flow. The evidence presented shows the differentiated cytotype of L. nigricans to be quite distinct from other L. nigricans accessions, both phenetically and phylogenetically. This indicates that the differentiated cytotype of L. nigricans may constitute a new taxon. Discriminant function analysis reveals that isozymes may be useful in validating species classification.     

Organelle based molecular analyses of the genetic relatedness of cultivated alfalfa (Medicago sativa L) to Medicago edgeworthii Sirjaev, and Medicago ruthenica (L.) Ledebour

Medicago edgeworthii Sirjaev and M. ruthenica (L.) Ledebour are allogamous, diploid (2n = 2x = 16) perennials with flat pods.Medicago edgeworthii is indigenous to the Himalayas and alpine areas west to Afghanistan, and Medicago ruthenica is found in Siberia, Mongolia, and Manchuria on open hillsides and mixed grass steppes. Because both species have a remarkable ability to survive extreme cold and poor soils, the possibility of hybridizing them with alfalfa (M. sativa L.) is being investigated. The objective of this research was to conduct an organelle based molecular assessment of the genetic relatedness of cultivated alfalfa (2n = 4x = 32) to M. edgeworthii and M. ruthenica. A hypervariable, intergenic region of cpDNA was amplified, and mtDNA was amplified with two primer pairs developed from soybean (Glycine max L.) mtDNA sequences. Mean Nei and Li genetic distances (GDs) between alfalfa and M. edgeworthii and alfalfa and M. ruthenica were 0.56 and 0.48 (mtDNA), and 0.33 and 0.30 (cpDNA), respectively. Intra specific GDs were 0.37 (mtDNA) and 0.25 (cpDNA) for M. edgeworthii; 0.42 (mtDNA) and 0.15 (cpDNA) for M. ruthenica; and 0 = 0.50 (mtDNA) and 0 = 0.23 (cpDNA) for alfalfa. Cluster analyses grouped someM. edgeworthii and M. ruthenica entries with alfalfa entries. There is some chance that alfalfa and M. edgeworthii entries which clustered closely could be hybridized; chances of alfalfa × M. ruthenica hybridizations appear to be more problematic. This revised version was published online in July 2006 with corrections to the Cover Date.     

Nematode resistance in bananas: screening results on some wild and cultivated accessions of Musa spp.

Bananas cultivated for export all belong to Cavendish cultivars and are all recognized as very susceptible to nematodes, particularly to the burrowing nematode Radopholus similis and the lesion nematode Pratylenchus coffeae. Even if there have been many changes in the management of banana nematodes in large commercial banana plantations, chemical control still remains most often the last resort method to manage the nematodes, although the number of registered products is definitely declining. Therefore, nematode control though genetic improvement is gaining new interest worldwide. In this study, 55 banana accessions mostly diploids from the Musa acuminata genome group (AA) but including some triploid accessions (AAA), some diploids of the Musa balbisiana genome group (BB) and some interspecific hybrids (AAB, AB) were evaluated for resistance to four nematode species R. similis, P. coffeae, Meloidogyne incognita and M. arenaria. These experiments were conducted in a growth chamber under controlled conditions. All banana accessions were susceptible to nematode species, although many different levels of susceptibility were detected. This study confirmed the good resistance status to R. similis of some cultivars from the Pisang jari buaya and Pisang batuau subgroups and the partial resistance of 17 diploid accessions significantly different from the susceptible reference cv. Grande Naine. This study also showed that 12 diploid accessions exhibited a partial resistance to P. coffeae, including some usual or potential genitors belonging to the wild diploids subspecies burmannica (cvs. Long Tavoy 1 and 2) and burmannicoides (cv. Calcutta 4). No source of resistance to Meloidogyne spp. was found. These screening results, combining for the first time four nematode species, are discussed within the scope of banana breeding in order to produce parental diploid lines with single or combined nematode resistances and further develop triploids that can substitute existing susceptible commercial cultivars.     

The myrobalan (<Emphasis Type="Italic">Prunus cerasifera</Emphasis> L.): a useful diploid model for studying the molecular genetics of self-incompatibility in plums

A series of PCR methods were used to detect S-RNase alleles and SFB alleles and to determine S-genotypes in 25 accessions of myrobalan (Prunus cerasifera L.). Firstly, primers flanking the polymorphic second intron were used to identify S-RNases in agarose gels. These primers amplified one or two bands per accession in 25 accessions. Then consensus primers were designed for amplifying the polymorphic first intron, unique to Prunus S-RNases, for automated fluorescent detection. Each accession produced one or two peaks. New primers were then developed to amplify the intron in the SFB gene, for detection by fluorescence. Cross-referencing PCR bands and peaks indicated 15 S-alleles were present in the 25 accessions. Cloning, sequencing and comparison with published data indicated that the amplified products were S-RNase alleles. Sequence information was used to design primers specific for each S-RNase. Full and consistent S-genotypes were obtained by cross-comparing PCR data for 23 of the 25 accessions, and two accessions appeared to have a single allele. Pollen-tube microscopy indicated function of some but not all of the S-alleles sequenced.     

PCR-RFLP analysis of chloroplast gene regions in Cajanus (Leguminosae) and allied genera

Twenty-eight species belonging to five genera of the sub tribe Cajaninae; viz., Cajanus(15 species), Rhynchosia (10 species), Dunbaria, Flemingia and Paracalyx were analysed for variations in four chloroplast generegions, rbcL, trnS-psbC, 16S and trnL-UAA. The four chloroplast gene regions were amplified with specific primers and subsequently digested with 15 restriction enzymes. Rhynchosia did not show any inter-specific differences in the four gene regions for any of the enzymes used. In Cajanus, the inter-specific PCR-RFLP profile of the four gene regions for all the enzymes were similar except for the Pst I digests of trnS-psbC gene region. At inter-generic level, rbcL gene region did not show polymorphism with any of the enzymes used; while in trnS-psbC gene region polymorphism was observed only with Pst I and Hae III digestions. Inter-generic PCR-RFLP of 16S did not reveal any variation. The trnL-UAA gene region had restriction site only forEcoR I in which Cajanus and Rhynchosia showed similar profiles while Dunbaria showed a different profile. The trnL-UAA gene region in Paracalyx and Flemingia did not have restriction site for this enzyme. Despite the analyses of four gene regions using 15 restriction enzymes, differentiation at genus level could not be obtained. These observations indicated limited divergence of the chloroplast genome among the four genera of the sub-tribe Cajaninae suggesting close relationships of the taxa. This revised version was published online in July 2006 with corrections to the Cover Date.     

Resistance to Rhopalosiphum padi (Homoptera: Aphididae) in Hordeum vulgare subsp. spontaneum and in hybrids with H. vulgare subsp. vulgare

Summary Accessions of Hordeum vulgare subsp. spontaneum, the progenitor of cultivated barley, were screened in field and glasshouse trials for resistance to the aphid Rhopalosiphum padi. A few selected lines were furthermore hybridized with a modern barley variety and the resulting populations evaluated. High levels of resistance were found among some of the spontaneum accessions resulting in lower aphid growth rates (maximum reduction 57%). Segregation patterns among siblings in F2 populations were continuous, indicating the presence of several genes with possibly additive effects. The usefulness of H. vulgare subsp. spontaneum for breeding aphid resistant barley is discussed.     

Diversity in selected wild and cultivated species of pigeonpea using RFLP of mtDNA

Diversity in 28 accessions representing 12 species of the genus, Cajanus arranged in 6 sections including 5 accessions of the cultivated species, C. cajan, and 4 species of the genus Rhyncosia available in the germplasm collection at ICRISAT was assessed using RFLP with maize mtDNA probes. Cluster analysis of the Southern blot hybridization data with 3 restriction enzymes – 3 probe combinations placed the genus Rhyncosia in a major group well separated from all the species belonging to the genus Cajanus. Within the genus Cajanus, the 4 accessions of C. platycarpus belonging to section Rhynchosoides formed a separate group in contrast to those in other sections of pigeonpea. In the section, Cajanus all the 5 accessions of C. cajan were grouped together and C. cajanifolius belonging to the same section was in a subgroup by itself closer to the main group. The four accessions of C. scarabaeoides, were together and the other species belonging to section Cantharospermum were in different subgroups. The intra-specific variation was seen even within accessions of certain pigeonpea wild species such as C. scarabaeoides, C. platycarpus, C. acutifolius, and even the cultivated species of C. cajan. This study suggests that RFLP of mtDNA can be used for the diversity analysis of pigeonpea and it gives some indications on the maternal lineage among the species. The variations in the mitochondrial DNA hybridization patterns also suggest the extensive rearrangement of the organelle genome among the Cajanus species. This revised version was published online in July 2006 with corrections to the Cover Date.     

Sources of resistance to diseases of sugar beet in related Beta germplasm: II. Soil-borne diseases

Between 580 and 700 accessions of related cultivated and wild species of the genus Beta were assessed for resistance to four soil-borne diseases of sugar beet: two seedling damping-off diseases caused by the fungi Aphanomyces cochlioides and Pythium ultimum and two diseases of more mature plants, Rhizoctonia root and crown rot, caused by the fungus R. solani, and Rhizomania, caused by Beet necrotic yellow vein virus (BNYVV), a furovirus transmitted by the plasmodiophorid Polymyxa betae. Analysis of resistance data (assessed on an international standardised 1–9 scale of Resistance Scores) indicated that the highest levels of resistance ({RS} 2) to A. cochlioides and P. ultimum were to be found amongst accessions of the more distantly related sections Corollinae (93% of accessions tested) and Procumbentes (10%), respectively; although useful levels could also be found in the more closely related, and sexually compatible, section Beta (1–6%). Resistance to Rhizoctonia was also found in section Beta (5–7%), depending on whether field or glasshouse tests were used, but there was little evidence of generally high levels of resistance to Rhizomania among accessions of this section. None of the accessions of sections Corollinae and Procumbentes exhibited any notable resistance to Rhizoctonia. However, all sections Procumbentes and some sections Corollinae (4%) accessions were highly resistant to Rhizomania. Individuals with high levels of resistance to Rhizomania were identified from within some section Beta and Corollinae accessions, in which there was evidence of segregation.     

Evolutionary studies of the soybean: the frequency and distribution of alleles among collections of Glycine Max and G. Soja of various origin

Summary Allelic frequencies at 10 loci common throughout the genus Glycine subgenus Soja were determined for 27 geographical area-species samples. The samples included 366 accessions of Glycine soja, the wild soybean, and 193 accessions of G. max, the domesticated soybean, from the USDA Soybean Germplasm Collection. The data indicate that the alleles for grey pubescence (t ₁), low seed coat peroxidase level (ep), and blunt pubescence tip (pb) probably arose as mutations during the domestication of G. max. The remaining seven loci studied(Fr, Pa ₁, Pa ₂, Fg ₁, Fg ₂, Fg ₃and Fg ₄)are polymorphic throughout the subgenus Soja; differences among collections of G. max and G. soja seem to be the result of differing selection pressures. A cluster analysis of allelic frequencies reveals two distinct groups within the subgenus corresponding to G. soja and G. max, Semi-wild accessions of G. max, while morphologically more similar to cultivated plants, clustered with samples of G. soja. The semi-wild accessions examined are thought to have arisen via hybridization between G. soja and G. max.Joint contribution: North Central Region, Agricultural Research, Science and Education Administration, U.S. Department of Agriculture, and Journal Paper No. J-9868 of the Iowa Agriculture and Home Economics Experiment Station, Ames, Iowa 50011; Project 2107.     

Avoidance of leaf rust fungi in wild relatives of cultivated cereals

Summary Avoidance of rust fungi that was based on poor appressorium induction was previously found in Hordeum chilense. In the present study 95 accessions of Triticeae were screened for avoidance of Puccinia hordei. The percentage of appressorium formation per germinated spore ranged from 6 to 90%. On none of the 41 accessions of Aegilops, Agropyron, Elymus, Secale, Thinopyrum or Triticum studied was the rate of appressorium formation lower than 25%. Lower rates of appressorium formation were, however, found on accessions of wild barley species Hordeum brachyantherum, H. marinum, H. parodii and H. secalinum. Its implications in cereal breeding are discussed.     

Assessment of genetic variability in grapefruits (Citrus paradisi Macf.) and pummelos (C. maxima (Burm.) Merr.) using RAPD and SSR markers

The genetic variability of 38 grapefruit (Citrus paradisi Macf.) and three pummelos (C. maxima (Burm.) Merr..) accessions was evaluated using RAPD, and single sequence repeat (SSR) analyses. Approximately49% of the 198 RAPD were polymorphic, and 4.6 alleles per SSR loci were identified. PIC values changed from 0.093 to 0.450. A UPGMA phenetic tree was constructed and two main grapefruit groups were identified. The grapefruit accessions `do Cabo' and `Siamesa-Filipinas'clustered very close to the pummelos in Group A. The Group B consisted of three sub-groups, which comprised all of the other grapefruit accessions. The majority of grapefruit accessions showed a narrow genetic base suggesting that the observed morphological polymorphism within the group must be associated with somatic mutations, which were not detected by these molecular markers. This revised version was published online in August 2006 with corrections to the Cover Date.     

DNA fingerprinting in soybean (Glycine max (L.) Merrill) with oligonucleotide probes for simple repetitive sequences

Summary Soybean DNA fingerprints were analyzed by digoxigenin-labeled oligonucleotide probes complementary to simple repetitive sequences. The clearest and most polymorphic patterns were obtained with (AAT)₆ as a probe, with which all 47 soybean cultivars tested could be distinguished. However, DNA fingerprints of individuals within cultivars showed the same pattern. Using (CT)₈, (GAA)₅ or (AAGG)₄ as probes, clear polymorphic patterns among cultivars and accessions in the subgenus Soja (Glycine max and Glycine soja) were not observed, while quite different patterns were found in accessions in the subgenus Glycine. The results suggest that G. max and G. soja are closer in their genome structure. DNA fingerprints of reciprocal crosses between cultivars and accessions in the subgenus Soja were similar, and contained bands of both parents. In an F₂ population from these crosses, such bands segregated in a Mendelian fashion.     

Identification of Resistance Genes Against Powdery Mildew in Four Accessions of Hordeum Vulgare SSP. Spontaneum

Summary Four newly detected accessions of wild barley (Hordeum vulgare ssp. spontaneum) resistant to powdery mildew caused by Blumeria graminis f. sp. hordei were studied with the aim of finding the number of genes/loci conferring the resistance of individual accessions, the type of inheritance of the genes and their relationships to the Mla locus. F₂ populations after crosses between the winter variety ‘Tiffany’ and four wild barley accessions and use of microsatellite DNA markers were focused on the identification of individual resistance genes/loci by means of their chromosomal locations. In PI466495, one locus conferring powdery mildew resistance was identified in highly significant linkage with the marker Bmac0213. This location is consistent with the known locus Mla on chromosome 1HS. In the other three accessions the resistance was determined by two independent loci. In PI466197, PI466297 and PI466461, one locus was identified on chromosome 1HS and three new loci were revealed on chromosomes 2HS (highly significant linkage with Bmac0134), 7HS (highly significant linkage with Bmag0021) and 7HL (significant linkage with EBmac0755). Our prospective aim is identification of further linked DNA markers and the exact location of the resistance genes on the barley chromosomes.     

Classification of Azolla spp., section Azolla

Summary Azolla accessions (section Azolla) from the germplasm collections of the International Rice Research Institute and Washington State University were fingerprinted and classified by enzyme electrophoresis and leaf trichome morphology. A. filiculoides was enzymatically distinctive and also reliably identified by its prominent one-celled trichomes. Neotropical accessions labelled as A. filiculoides proved to be members of other species. Two groups of isolates were designated A. rubra, but those from Japan were identified as A. filiculoides. The A. rubra of Australia-New Zealand was biochemically unique and possessed less protuberant trichomes than A. filiculoides. A. microphylla, A. mexicana, and A. caroliniana were phenetically similar, but a. microphylla was identifiable from the others in the banding patterns of certain enzymes. A. mexicana and A. caroliniana were closely related enzymatically. The two-celled leaf trichomes of these three species were similar in size and shape.